Figure 6: TERT regulates MYC half-life and ubiquitination.

Transcription

1 TERT or IgG as indicated. For the western blots, representative images of n= independent experiments are shown. Student s t-test was used, and * indicates p<.5 and # indicates a non-significant p-value, compared to sh/sicontrol. Figure 6: TERT regulates MYC half-life and ubiquitination. (A) and (B) Western blots showing MYC levels upon TERT or Terc knockdown following cycloheximide treatment (CHX) for indicated time points in P9 cells and EµMYC B cells. (C) and (D) Western blots in P9 and EmMYC B cells showing the levels of MYC, TERT and ACTIN upon TERT knockdown, with and without MG treatment ( µm, h). (E) Western blots showing levels of indicated proteins upon vector, TERT WT and TERT DN expression. (F) Coimmunoprecipitation between endogenous MYC and GSKβ in cells infected with shrna against control (-) or TERT(+) and immunoblotting with indicated antobodies. (G) 9T cells were transfected with HA-MYC, His-ubiquitin along with or shtert. 8h post transfection cells were harvested, immunoprecipitated with Ni-NTA beads, and western blotted with HA. Right panel shows the inputs. (H) In vivo MYC ubiquitination assay in cells transfected with control (-) or TERT (+) shrnas along with HA-Ub. Immunoprecipitation and immunoblots from lysates or immunoprecipitaed material were performed by using indicated antibodies. For the western blots, representative images of n= independent experiments are shown. Figure 7: Graphical model. TERT levels, which are limiting in reconstituting telomerase activity in normal cells, are upregulated by increased MYC in cancer cells. TERT, in turn, enhances MYC stability and function, thereby regulating its own levels and telomerase activity. This function of TERT does not require Terc, and is independent of its function at telomeres. Enhanced MYC stability and function in high TERT and high MYC cells, which is a consequence of this feed forward mechanism, leads to enhanced oncogenesis due to downstream targets of MYC, which are known to regulate aspects of cell cycle, proliferation and metabolism. Supplementary Figure : (A) Western blot of MYC and TERT in B-cells from EµMYC mice (n=) compared to wildtype mice (n=). (B) Viability of EµMYC B-cells following TERT knockdown using independent shrnas, which could be partially rescued by the overexpression of human TERT. (C)

2 Viability of wildtype and EµMYC B-cells following TERT knockdown. (D) TERT protein levels, as shown by Western blot (top), in primary lymphoma cells from EµMYC mice after silencing Terc (sh- Terc) (bottom). Values denote mean+s.d., n=. (E) Teloblot showing the length of telomeres in primary lymphoma cells from EµMYC mice after silencing Terc (shterc) or TERT (shtert). One way ANOVA with Tukey s Multiple Comparison Test was used for (B and C) and students t-test was used for (D) * indicates p<.5 vs., indicates p<. vs., ^ indicates non-significant p-value vs. and indicates p<. vs. shtert-a or shtert-b. For the western blots, representative images of n= independent experiments are shown. Supplementary Figure : (A) Teloblot showing of telomere lengths in tumors from EµMYC;TERT+/+ and EµMYC;TERT-/- mice (n= for each genotype). (B) Immunofluorescence staining for TRF (green) and γhax (red) in EµMYC;TERT+/+, and EµMYC;TERT-/- B cells The cell nuclei were counterstained with DAPI (blue). We did not observe any DNA damage foci (γhax staining) at telomeric regions. As B cells are small and round, the staining is less distinct. Representative images are shown, and cells isolated from mice of each genotype were analyzed (6x magnification). Supplementary Figure : (A) ChIP-seq binding profiles at selected promoters in P9 cells (B) and EµMYC;TERT+/+ and EµMYC;TERT-/- B cells. (C, D) Validation of selected ChIP seq targets in P9 cells with additional shrna constructs. These changes could be rescued by the overexpression of TERT WT or TERT DN. (E) Validation of selected ChIP seq targets in B cells isolated from -week old, pretumoral EµMYC;TERT+/+ and EµMYC;TERT-/- mice (n= for each genotype) using antibodies for MYC and Pol-II. The B-cells from EµMYC;TERT-/- showed lower enrichment of MYC and Pol-II at the promoters of selected MYC-regulated genes. (F) NF-Y ChIP was carried out in P9 cells infected with and shtert. Graphs show % Input for indicated target promoters. (G) Heatmap showing the validation of the expression of selected Myc-target genes in -week-old EµMYC;TERT+/+, EµMYC;TERT+/- and EµMYC;TERT-/- spleen B cells. (H-I) Validation of selected gene expression changes in P9 cells with indicated infections. X axis labels indicate rrna target genes and rdna regions respectively. (J) Validation of selected gene expression changes in EµMYC B cells. Values denote mean+s.d., n=. One way ANOVA with Tukey s Multiple Comparison Test was used for (C, D, H, I and J) and students t-test was used for (E) and (F) * indicates p<.5 vs., indicates p<. vs., ^ indicates a non-significant p-value vs., # indicates p<.5 vs. shtert (for C, D, H, I) or shtert-a or shtert-b (for J), indicates p<. vs. shtert (for C, D, H, I) or 5

3 shtert-a or shtert-b (for J) indicates a non-significant p-value vs. shtert (for C, D, H, I) or shtert-a or shtert-b (for J). Supplementary Figure : (A-B) Gene expression analysis in P9 cells with knockdown of TERT and Terc under high MYC (cells grown in no tetracycline, no estradiol) and low MYC conditions [cells grown with tetracycline (. µg/ml) and β-estradiol ( µm)]. Values denote mean+s.d., n=. Students t-test was used and * indicates p<.5. Supplementary Figure 5: (A) VA cells were transfected with the indicated expression vectors. 8 h post transfection, the samples were harvested and processed for quantitative RT-PCR. The graph shows relative TERT expression. ACTIN was used as the reference gene. (Left panel). VA cells were transfected with the indicated expression vectors. 8 h post transfection, the samples were harvested and processed for western blotting of MYC and ACTIN (right panel). (B) MYC was immunoprecipitated from MIHA cells and the eluates were western blotted for indicated proteins. (C) Co-Immunoprecipitation between endogenoeus MYC and TERT in the P9 cells with, shtert or shterc. (D) Schematic showing the various domains of MYC. (E) 9T cells were transfected with FLAG-tagged human wild-type or MYC mutants deleted in amino acid residues -, 9-5, -8, -, -8 and 8-8, or vector control. Bottom Panel shows western blots indicating input proteins and top panel shows western blot analysis of indicated lysates immunoprecipitated with Flag or TERT antibody. Red arterisk indicates no interaction. (F) Left panel: Bacterially expressed GST, pgext-myc (-8 aa) or pgexx-myc (98-9 aa) proteins were incubated with whole cell lysates from Hela with control or TERT overexpression. Following incubation, the mixture was immunoprecipitated with GST antobody and immunoblotted (IB) with TERT. Right Panel: Coomasie Blue staining of Purified MYC proteins (- 8 aa) and (98-9 aa) along with GST expressed from a control vector (GST). (G) 9T cells were transfected with indicated proteins and checked for gene expression. (H) 9T cells were transfected with FLAG-tagged human wild-type, dominant negative or TERT mutants deleted in amino acid residues - 6, -7, -95, 5-95 or Flag vector control. The bottom panel shows western blots indicating input proteins and top panel shows western blot analysis of indicated lysates immunoprecipitated with Flag antibody. For the western blots, representative images of n= independent experiments are shown. One way ANOVA with Tukey s Multiple Comparison Test was used, and * indicates p<.5 vs. Vector, indicates p<. vs. Vector, ^ indicates a non-significant p-value vs. Vector. 6

4 Supplementary Figure 6: (A) Western blots showing MYC levels following cycloheximide treatment (CHX) for the indicated time points in P9 cells infected with, shtert and. (B) Western blots showing MYC levels upon cycloheximide treatment (CHX) for indicated time points in 9T cells transfected with Flag-MYC T58A and or shtert. (C) Gene expression analysis in cells transfected with the indicated plasmids. Target genes are indicated on x-axis. (D) In vivo MYC ubiquitination assay in cells transfected with shtert and Flag MYC along with HA-Ub. (E) 9T cells were transfected with Flag-MYC Δ -, His-ubiquitin along with or shtert. 8 h post transfection, the cells were harvested, immunoprecipitated with Ni-NTA beads (IP:HIS), and western blotted with Flag. The right panel shows the inputs. One way ANOVA with Tukey s Multiple Comparison Test was used. * indicates p<.5 vs., indicates p<. vs., ^ indicates not significant vs., indicates p<. vs. shtert + vector. For the western blots, representative images of n= independent experiments are shown. Supplementary Figure 7: Vector and TERT overexpressing P9 cells were transfected with sicontrol or sibrg for 8h. A) panel shows relative Brg expession. B) panel shows gene expression in the various transfected conditions. SUPPLEMENTARY TABLES Supplementary Table. shrna sequences. Gene shtert-a shtert-b shtert shtert shtert (targets UTR) shrna sequence TTCTGTCATCTACAAACTCGAGTTTGTAGATGACAGAA ACTATGAGCGGACAAACTCGAGTTTGTCCGCTCATAGT CATTTCATCAGCAAGTTTGA GAAGAGTGTCTGGAGCAAGTT TGAGGCCTGAGTGAGTGTTTG Supplementary Table. qpcr primers for gene expression analysis. 7

5 Human Gene Forward Primer Reverse Primer PKM TGCGAGCCTCAAGTCACTCCAC TCACGGCACAGGAACAACACG LDHA GGCCTGTGCCATCAGTATCT ACCAGCTTGGAGTTTGCAGT ALDOC AAATTGGGGTGGAAAACACA ACCCTTGTCAACCTTGATGC HK TCTATGCCATCCCTGAGGAC TCTCTGCCTTCCACTCCACT GAPDH GGCGATGCTGGCGCTGAGTA ACAGTTTCCCGGAGGGGCCA c-myc ACGGCCGACCAGCTGGAGAT TCGGGCTGCCGCTGTCTTTG TERT CCAAGTTCCTGGACTGGCTGA TTCCCGATGCTGCCTGAC CYCLIN D AGTGCGTGCAGAAGGACATC GTTGCAGATGGGACTTCGGA CYCLIN D GTGCTGCGAAGTGGAAACCATC GACCTCCTTCTGCACACATTTGA CDK GCCTTCCCATCAGCACAGTTC GTCTACATGCTCAAACACCAGGG eifa ATGGGACCTTGTTTGCCTGG CCACGTTGCCAGGACAGTAT 5s rrna GAACGGTGGTGTGTCGTTC GCGTCTCGTCTCGTCTCACT 5.8s rrna ACTCGGCTCGTGCGTC GCGACGCTCAGACAGG BRG ACCAGATGCACAAGCCCATG GCTGGAACTGGACTAGAGGC β-actin GCCAACCGCGAGAAGATGA CCATCACGATGCCAGTGGTA MOUSE Gene Forward primer Reverse primer TERT GGATTGCCACTGGCTCCG TGCCTGACCTCCTCTTGTGAC 5s rrna CGCCCGCGTGTTGGTCTTCT CCCAAAACCCCGACGAGCCC 5.8s rrna ACTCGGCTCGTGCGTC GCGACGCTCAGACAGG 8s rrna GCCTGCTGCCTTCCTTGG GATGGTAGTCGCCGTGCC 8s rrna AGAGGTAAACGGGTGGGGTC GGGGTCGGGAGGAACGG GAPDH CATCTTCTTGTGCAGTGCCAG GGCAACAATCTCCACTTTGCC ENO GAGCGAGAAGTCCTGCAACT CTGCGATGAAAGTGTCCTCA 8

6 LDHA AGACAAACTCAAGGGCGAGA GCGGTGATAATGACCAGCTT PKM TGACACCTTCCTGGAACACA TTCAGCATCTCCACAGATCG EXOSC GCTCCTCTCATCCATGAACCT CAGCTCCATCCGAGAACACA GART CGTCATTGCTGGAATTGCT TTGGGCATCTCTGCTGTCT CCT7 CCAGTTATCCTGTTGAAAGAGGG GACCCAGGGTGGTTCTTACA MCM6 GCTGTTCCTAGACTTCCTGGA CAACCAGCGTGTTTCTCTCAG MYBBPA GCCGCGAGTTCTTGGACTT ATCTCCGAATCATTGGGCCTT β-actin CAGCTTCTTTGCAGCTCCTT CAGCTTCTTTGCAGCTCCTT Supplementary Table. ChIP primers. Human Gene Forward Primer Reverse Primer LDHA GGAGGGCAGCACCTTACTTA AGGGGCCTTAAGTGGAACAG CDK GAGCGACCCTTCCATAACCA GGGCTGGCGTGAGGTAAGT HK GATTGCCTCGCATCTGC AGCCACGATTCTCTCCACG PKM TTTGGAGGAGTCTGCAAAGG CACTCCCAAGGACCAAGTGT TERT GCGCGAGTTTCAGGCAG CACCTCGCGGTAGTGGCT rdna (H) GGCGGTTTGAGTGAGACGAGA ACGTGCGCTCACCGAGAGCAG rdna (H) CGACGACCCATTCGAACGTCT CTCTCCGGAATCGAACCCTGA ZNF77 GCACCCCTGATTGGAGAAT TCAGAGACGTTAATCCGAAGC JMJDB CTGAGGCTCCCTTGTCAATC CGGCTCTGTTCTCATTGGAG RUVBL GAGGAGGAGGGTGGGATAAA GGCCGATTGCAAAATTCTTA Mouse Gene Forward Primer Reverse Primer BXDC GATCCCTTAGCGAATGCACG GGCTTCCCGGAAGTAATCG 9

7 GART HSPD MYBBPA ACCTCCATCAAGGCCGTGC CGCCACACGTGATGAAACC AGCCCACGTGTTTGGCTC GTGACTGGCGAAAAGCCCAC TCGGCTCACTTGTCCAGGC GGGCTCTTCATCTCCGCC Supplementary Table. Statistical analysis.

8 Sta$s$cs for Figure D One- way analysis of variance P value P<. * Are means signif. different? (P <.5) Yes Number of groups 7 F 8. R squared.685 ANOVA Table SS df MS Treatment (between columns). 6.5 Residual (within columns) Total.95 9 DunneW's Mul$ple Comparison Test Mean Diff. q P value 95% CI of diff Control vs shtert- A P <.5.85 to.89 Control vs shtert- B P <..5 to. Control vs shterc P > to.68 Control vs TERT P > to.79 Control vs TERT; shtert- A.58.8 P > to.7 Control vs TERT; shtert- B P > to.

9 Sta$s$cs for Fig SB F 7. P value <. Are differences among means statistically significant Yes R square.9769 F (DFn, DFd).7 (5, 5) P value.898 ns Significantly different standard deviations? (P <.5)No Bartlett's statistic (corrected) P value Significantly different standard deviations? (P <.5) ANOVA table SS DF MS F (DFn, DFdP value Treatment (between columns) F (5, 5) = P <. Residual (within columns). 5.7 Total.78 Number of treatments (columns) 6 Number of values (total) Number of families Number of comparisons per family 5 Alpha.5 Tukey's multiple comparisons test Mean Diff. 95% CI of diff. Significant? Summary vs. shtert-a.5.8 to.66 Yes vs. shtert-b to.878 Yes vs. +TERT to.7 No ns vs. shtert-a +TERT to.9 Yes * vs. shtert-b +TERT to.565 Yes shtert-a vs. shtert-b.8.95 to.5 Yes * shtert-a vs. +TERT to -.5 Yes shtert-a vs. shtert-a +TERT to -. Yes shtert-a vs. shtert-b +TERT to -.66 Yes shtert-b vs. +TERT to -.67 Yes shtert-b vs. shtert-a +TERT to -.5 Yes shtert-b vs. shtert-b +TERT to -.5 Yes +TERT vs. shtert-a +TERT..8 to.6 Yes +TERT vs. shtert-b +TERT to.57 Yes shtert-a +TERT vs. shtert-b +TERT.66.7 to.5 Yes * Symbol Meaning ns P >.5 * P.5 P. * P. P.

10 Sta$s$cs for Figure SC F 59. P value <. Are differences among means statistically significant?yes R square.9779 F (DFn, DFd) (5, 8) P value. * Significantly different standard deviations? (P <.5) Yes Bartlett's statistic (corrected) +infinity P value <. Significantly different standard deviations? (P <.5) Yes ANOVA table SS DF MS F (DFn, DFd) P value Treatment (between columns) F (5, 8) = 59. P <. Residual (within columns) Total.9 Number of treatments (columns) 6 Number of values (total) Number of families Number of comparisons per family 5 Alpha.5 Tukey's multiple comparisons test Mean Diff. 95% CI of diff. Significant? Summary Wildtype- vs. Wildtype- shtert-a to.5 No ns Wildtype- vs. Wildtype- shtert-b..65 to.8 Yes * Wildtype- vs. EuMYC- -.8 to.8 No ns Wildtype- vs. EuMYC- shtert-a.5.9 to.6569 Yes Wildtype- vs. EuMYC- shtert-b to.877 Yes Wildtype- shtert-a vs. Wildtype- shtert-b to.5 No ns Wildtype- shtert-a vs. EuMYC to.57 No ns Wildtype- shtert-a vs. EuMYC- shtert-a.59.8 to.5657 Yes Wildtype- shtert-a vs. EuMYC- shtert-b to.78 Yes Wildtype- shtert-b vs. EuMYC to -.65 Yes * Wildtype- shtert-b vs. EuMYC- shtert-a..9 to.57 Yes Wildtype- shtert-b vs. EuMYC- shtert-b to.757 Yes EuMYC- vs. EuMYC- shtert-a.5.9 to.6569 Yes EuMYC- vs. EuMYC- shtert-b to.877 Yes EuMYC- shtert-a vs. EuMYC- shtert-b.8.9 to.86 Yes * Symbol Meaning ns P >.5 * P.5 P. * P. P.

11 Sta$s$cs for Figure D WBC One- way analysis of variance P value.87 Are means signif. different? (P <.5) Yes Number of groups F 5.6 R squared.668 BartleT's test for equal variances BartleT's sta$s$c (corrected) 6.79 P value P<. * Do the variances differ signif. (P <.5) Yes ANOVA Table SS df MS Treatment (between columns) Residual (within columns) Total 67 5 DunneT's Mul$ple Comparison Test Mean Diff. q P value 95% CI of diff EuMYC; TERT+/+ vs EuMYC; TERT+/-..8 P < to.59 EuMYC; TERT+/+ vs EuMYC; TERT- / P <..76 to 7. Tumor One- way analysis of variance P value.9 Are means signif. different? (P <.5) Yes Number of groups F 5.8 R squared.89 BartleT's test for equal variances BartleT's sta$s$c (corrected).7 P value P<. * Do the variances differ signif. (P <.5) Yes ANOVA Table SS df MS Treatment (between columns) Residual (within columns) Total.5 8 DunneT's Mul$ple Comparison Test Mean Diff. q P value 95% CI of diff EuMYC; TERT+/+ vs EuMYC; TERT+/-.8.5 P <.5. to. EuMYC; TERT+/+ vs EuMYC; TERT- / P <..58 to.5 Spleen (% body weight) One- way analysis of variance P value.67 Are means signif. different? (P <.5) Yes Number of groups F R squared.957 BartleT's test for equal variances BartleT's sta$s$c (corrected).7 P value P<. * Do the variances differ signif. (P <.5) Yes ANOVA Table SS df MS Treatment (between columns)..6 Residual (within columns) Total.6 8 DunneT's Mul$ple Comparison Test Mean Diff. q P value 95% CI of diff EuMYC; TERT+/+ vs EuMYC; TERT+/-.7.5 P > to.977 EuMYC; TERT+/+ vs EuMYC; TERT- / P <..9 to.

12 Sta$s$cs for Figure SC Cyclin D F 59. P value <. Are differences among means statistically signyes R square.977 F (DFn, DFd).7 (5, ) P value.55 * Significantly different standard deviations? (P <Yes Bartlett's statistic (corrected).8 P value.9 * Significantly different standard deviations? (P <Yes ANOVA table SS DF MS F (DFn, DFdP value Treatment (between columns) F (5, ) = P <. Residual (within columns) Total Number of treatments (columns) 6 Number of values (total) 6 Number of families Number of comparisons per family 5 Alpha.5 Tukey's multiple comparisons test Mean Diff. 95% CI of d Significant? Summary vs. shtert.7.6 to. Yes vs. shtert.9.76 to. Yes vs. shtert.6.7 to. Yes vs. shtert + TERT WT to No ns vs. shtert + TERT DN to No ns shtert vs. shtert to Yes * shtert vs. shtert to -Yes shtert vs. shtert + TERT WT to -Yes shtert vs. shtert + TERT DN to -Yes shtert vs. shtert to Yes shtert vs. shtert + TERT WT to -Yes shtert vs. shtert + TERT DN to -Yes shtert vs. shtert + TERT WT to -Yes shtert vs. shtert + TERT DN to -Yes shtert + TERT WT vs. shtert + TERT D to No ns Nucleolin F 75 P value <. Are differences among means statistically signyes R square.9957 F (DFn, DFd).8 (5, ) P value.65 Significantly different standard deviations? (P <Yes Bartlett's statistic (corrected).8 P value.9 * Significantly different standard deviations? (P <Yes ANOVA table SS DF MS F (DFn, DFdP value Treatment (between columns) F (5, ) = P <. Residual (within columns) Total Number of treatments (columns) 6

13 Number of values (total) 6 Number of families Number of comparisons per family 5 Alpha.5 Tukey's multiple comparisons test Mean Diff. 95% CI of d Significant? Summary vs. shtert to. Yes vs. shtert to. Yes vs. shtert to. Yes vs. shtert + TERT WT tono ns vs. shtert + TERT DN.9.88 to. Yes shtert vs. shtert to -Yes shtert vs. shtert to -Yes shtert vs. shtert + TERT WT to -Yes shtert vs. shtert + TERT DN to -Yes shtert vs. shtert to -Yes shtert vs. shtert + TERT WT to -Yes shtert vs. shtert + TERT DN to -Yes shtert vs. shtert + TERT WT to -Yes shtert vs. shtert + TERT DN to -Yes shtert + TERT WT vs. shtert + TERT D to. Yes eifa F 9.6 P value <. Are differences among means statistically signyes R square.87 F (DFn, DFd).66 (5, ) P value.8 * Significantly different standard deviations? (P <Yes Bartlett's statistic (corrected) 5. P value.99 Significantly different standard deviations? (P <Yes ANOVA table SS DF MS F (DFn, DFdP value Treatment (between columns) F (5, ) = P <. Residual (within columns) Total.55 5 Number of treatments (columns) 6 Number of values (total) 6 Number of families Number of comparisons per family 5 Alpha.5 Tukey's multiple comparisons test Mean Diff. 95% CI of d Significant? Summary vs. shtert.6.8 to. Yes vs. shtert..685 to Yes vs. shtert to Yes * vs. shtert + TERT WT to No ns vs. shtert + TERT DN to No ns shtert vs. shtert to No ns shtert vs. shtert to -Yes shtert vs. shtert + TERT WT to -Yes shtert vs. shtert + TERT DN to -Yes shtert vs. shtert to No ns shtert vs. shtert + TERT WT to -Yes shtert vs. shtert + TERT DN to -Yes shtert vs. shtert + TERT WT to -Yes * shtert vs. shtert + TERT DN to -Yes * shtert + TERT WT vs. shtert + TERT D to No ns Symbol Meaning ns P >.5 * P.5

14 P. * P. P.

15 Sta$s$cs For Figure SD H F.5 P value <. Are differences among means statistically signyes R square.66 F (DFn, DFd) 9.75 (5, ) P value <. Significantly different standard deviations? (P <Yes Bartlett's statistic (corrected) 5.59 P value <. Significantly different standard deviations? (P <Yes ANOVA table SS DF MS F (DFn, DFd) P value Treatment (between columns) F (5, ) =.5 P <. Residual (within columns).6.57 Total.75 5 Number of treatments (columns) 6 Number of values (total) 6 Number of families Number of comparisons per family 5 Alpha.5 Tukey's multiple comparisons test Mean Diff. 95% CI of diff. Significant? Summary vs. shtert.6.67 to.99yes * vs. shtert to.yes * vs. shtert to.8yes * vs. shtert + TERT WT. -.5 to.6no ns vs. shtert + TERT DN. -. to.no ns shtert vs. shtert. -. to.no ns shtert vs. shtert to.9no ns shtert vs. shtert + TERT WT to.7no ns shtert vs. shtert + TERT DN to -.Yes * shtert vs. shtert to.6no ns shtert vs. shtert + TERT WT to.no ns shtert vs. shtert + TERT DN to -.Yes * shtert vs. shtert + TERT WT to.5no ns shtert vs. shtert + TERT DN to -.Yes * shtert + TERT WT vs. shtert + TERT D to.8no ns H F P value <. Are differences among means statistically signyes R square.99 F (DFn, DFd) 6.67 (5, ) P value. * Significantly different standard deviations? (P <Yes Bartlett's statistic (corrected). P value.7 * Significantly different standard deviations? (P <Yes ANOVA table SS DF MS F (DFn, DFd) P value Treatment (between columns) F (5, ) =. P <. Residual (within columns) E-6 Total.8 5 Number of treatments (columns) 6 Number of values (total) 6 Number of families Number of comparisons per family 5 Alpha.5

16 Tukey's multiple comparisons test Mean Diff. 95% CI of diff. Significant? Summary vs. shtert.5.68 to.6yes vs. shtert.89.8 to.yes vs. shtert to.yes vs. shtert + TERT WT.6.8 to.yes * vs. shtert + TERT DN.8.79 to.yes shtert vs. shtert to.no ns shtert vs. shtert to -.Yes shtert vs. shtert + TERT WT to -.Yes shtert vs. shtert + TERT DN to -.Yes shtert vs. shtert to -.Yes shtert vs. shtert + TERT WT to -.Yes shtert vs. shtert + TERT DN to -.Yes shtert vs. shtert + TERT WT to -.Yes shtert vs. shtert + TERT DN to.no ns shtert + TERT WT vs. shtert + TERT D to.yes * Symbol Meaning ns P >.5 * P.5 P. * P. P.

17 Sa#s#cs for Figure SH 5s F 9.9 P value <. Are differences among means statistically significayes R square.98 F (DFn, DFd).795 (6, 5) P value.5 * Significantly different standard deviations? (P <. Yes Bartlett's statistic (corrected).7 P value.58 ns Significantly different standard deviations? (P <. No ANOVA table SS DF MS F (DFn, DFdP value Treatment (between columns) F (6, 5) = P <. Residual (within columns) Total 5.98 Number of treatments (columns) 7 Number of values (total) Number of families Number of comparisons per family Alpha.5 Tukey's multiple comparisons test Mean Diff. 95% CI of diff. Significant? Summary vs. shterc.6. to.7 Yes * vs. shtert to.87 Yes vs. shtert to.85 Yes vs. shtert.89.6 to.5 Yes vs. shtert + TERT WT to -.7 Yes vs. shtert + TERT DN to.595no ns shterc vs. shtert.6.58 to.76 Yes shterc vs. shtert to.76 Yes shterc vs. shtert.5.68 to.88 Yes shterc vs. shtert + TERT WT to -.9 Yes shterc vs. shtert + TERT DN to -.5 Yes * shtert vs. shtert to.no ns shtert vs. shtert to -.55 Yes shtert vs. shtert + TERT WT to Yes shtert vs. shtert + TERT DN to Yes shtert vs. shtert to -.95 Yes shtert vs. shtert + TERT WT to Yes shtert vs. shtert + TERT DN to -.77 Yes shtert vs. shtert + TERT WT to -.7 Yes shtert vs. shtert + TERT DN to -.77 Yes shtert + TERT WT vs. shtert + TERT DN to.5 Yes

18 5.8s F.9 P value <. Are differences among means statistically significayes R square.968 F (DFn, DFd).689 (6, 5) P value.98 * Significantly different standard deviations? (P <. Yes Bartlett's statistic (corrected).6 P value <. Significantly different standard deviations? (P <. Yes ANOVA table SS DF MS F (DFn, DFdP value Treatment (between columns) F (6, 5) = P <. Residual (within columns) Total.8 Number of treatments (columns) 7 Number of values (total) Number of families Number of comparisons per family Alpha.5 Tukey's multiple comparisons test Mean Diff. 95% CI of diff. Significant? Summary vs. shterc to.98no ns vs. shtert to.856 Yes vs. shtert to.89 Yes vs. shtert to.57 Yes vs. shtert + TERT WT to -.79 Yes vs. shtert + TERT DN to.yes * shterc vs. shtert.56.8 to.765 Yes shterc vs. shtert.55.9 to.6996 Yes shterc vs. shtert..59 to.676 Yes shterc vs. shtert + TERT WT to -.57 Yes shterc vs. shtert + TERT DN to.77 No ns shtert vs. shtert to.7 No ns shtert vs. shtert to -.97 Yes shtert vs. shtert + TERT WT to Yes shtert vs. shtert + TERT DN to Yes shtert vs. shtert to Yes shtert vs. shtert + TERT WT to -.87 Yes shtert vs. shtert + TERT DN to -.76 Yes shtert vs. shtert + TERT WT to -.77 Yes shtert vs. shtert + TERT DN to -.86 Yes shtert + TERT WT vs. shtert + TERT DN..778 to.586 Yes Symbol Meaning ns P >.5 * P.5 P. * P. P.

19 Sta$s$cs for Figure SI Cyclin D F 55.9 P value <. Are differences among means statistically significant? (P <.5) Yes R square.9896 F (DFn, DFd) 5.59 (6, 5) P value.8 * Significantly different standard deviations? (P <.5) Yes Bartlett's statistic (corrected) 8.9 P value.5 Significantly different standard deviations? (P <.5) Yes ANOVA table SS DF MS F (DFn, DFdP value Treatment (between columns) F (6, 5) = 5P <. Residual (within columns) Total 5. Number of treatments (columns) 7 Number of values (total) Number of families Number of comparisons per family Alpha.5 Tukey's multiple comparisons test Mean Diff. 95% CI of d Significant? Summary vs. shterc to No ns vs. shtert to. Yes vs. shtert to. Yes vs. shtert to. Yes vs. shtert + TERT WT to -.Yes vs. shtert + TERT DN to -.Yes shterc vs. shtert..5 to. Yes shterc vs. shtert to. Yes shterc vs. shtert.5.97 to. Yes shterc vs. shtert + TERT WT to -.Yes shterc vs. shtert + TERT DN to -.Yes shtert vs. shtert.8 -. to No ns shtert vs. shtert.8 -. to No ns shtert vs. shtert + TERT WT to -.Yes shtert vs. shtert + TERT DN to -.Yes shtert vs. shtert to No ns shtert vs. shtert + TERT WT to -.Yes shtert vs. shtert + TERT DN to -.Yes shtert vs. shtert + TERT WT to -.Yes shtert vs. shtert + TERT DN to -.Yes shtert + TERT WT vs. shtert + TERT DN to. Yes

20 eifa F.5 P value <. Are differences among means statistically significant? (P <.5) Yes R square.97 F (DFn, DFd).7 (6, 5) P value.58 Significantly different standard deviations? (P <.5) Yes Bartlett's statistic (corrected) 8.5 P value <. Significantly different standard deviations? (P <.5) Yes ANOVA table SS DF MS F (DFn, DFdP value Treatment (between columns) F (6, 5) = P <. Residual (within columns) Total 9.75 Number of treatments (columns) 7 Number of values (total) Number of families Number of comparisons per family Alpha.5 Tukey's multiple comparisons test Mean Diff. 95% CI of d Significant? Summary vs. shterc.9.98 to Yes vs. shtert to. Yes vs. shtert to. Yes vs. shtert to. Yes vs. shtert + TERT WT to -Yes vs. shtert + TERT DN to No ns shterc vs. shtert.6.6 to. Yes shterc vs. shtert to. Yes shterc vs. shtert to. Yes shterc vs. shtert + TERT WT to -Yes shterc vs. shtert + TERT DN to -Yes shtert vs. shtert to No ns shtert vs. shtert to No ns shtert vs. shtert + TERT WT to -.Yes shtert vs. shtert + TERT DN to -Yes shtert vs. shtert to No ns shtert vs. shtert + TERT WT to -.Yes shtert vs. shtert + TERT DN to -Yes shtert vs. shtert + TERT WT to -.Yes shtert vs. shtert + TERT DN to -Yes shtert + TERT WT vs. shtert + TERT DN.5.68 to. Yes

21 Nucleolin F 6. P value <. Are differences among means statistically significant? (P <.5) Yes R square.9876 F (DFn, DFd). (6, 5) P value.89 Significantly different standard deviations? (P <.5) Yes Bartlett's statistic (corrected) 5. P value. * Significantly different standard deviations? (P <.5) Yes ANOVA table SS DF MS F (DFn, DFdP value Treatment (between columns) F (6, 5) = P <. Residual (within columns) Total Number of treatments (columns) 7 Number of values (total) Number of families Number of comparisons per family Alpha.5 Tukey's multiple comparisons test Mean Diff. 95% CI of d Significant? Summary vs. shterc to Yes vs. shtert to. Yes vs. shtert to. Yes vs. shtert to. Yes vs. shtert + TERT WT to -Yes vs. shtert + TERT DN.8.7 to. Yes shterc vs. shtert to. Yes shterc vs. shtert to. Yes shterc vs. shtert.59.5 to. Yes shterc vs. shtert + TERT WT to -Yes shterc vs. shtert + TERT DN.6.86 to Yes shtert vs. shtert to No ns shtert vs. shtert to No ns shtert vs. shtert + TERT WT to -.Yes shtert vs. shtert + TERT DN to -Yes shtert vs. shtert to No ns shtert vs. shtert + TERT WT to -.Yes shtert vs. shtert + TERT DN to -Yes shtert vs. shtert + TERT WT to -Yes shtert vs. shtert + TERT DN to -Yes shtert + TERT WT vs. shtert + TERT DN to. Yes Symbol Meaning ns P >.5 * P.5 P. * P. P.

22 Sta$s$cs for Figure SJ ENO F.7 P value <. Are differences among means statistically significant? (P <.5) Yes R square.899 F (DFn, DFd).9 (, ) P value.8 ns Significantly different standard deviations? (P <.5) No Bartlett's statistic (corrected) P value Significantly different standard deviations? (P <.5) ANOVA table SS DF MS F (DFn, DFd) P value Treatment (between columns) F (, ) =.7 P <. Residual (within columns) Total.68 Number of treatments (columns) 5 Number of values (total) 5 Number of families Number of comparisons per family Alpha.5 Tukey's multiple comparisons test Mean Diff. 95% CI of diff. Significant? Summary vs. shtert-a to.6776 No ns vs. shtert-b.6.55 to.95 Yes vs. shtert-a +TERT to.56 No ns vs. shtert-b +TERT to.59 No ns shtert-a vs. shtert-b to.65 No ns shtert-a vs. shtert-a +TERT to -.76 Yes shtert-a vs. shtert-b +TERT to -.9 Yes shtert-b vs. shtert-a +TERT to -. Yes * shtert-b vs. shtert-b +TERT to -.75 Yes * shtert-a +TERT vs. shtert-b +TERT to.96 No ns GAPDH F 5.59 P value <. Are differences among means statistically significant? (P <.5) Yes R square.9 F (DFn, DFd).65 (, ) P value.67 ns Significantly different standard deviations? (P <.5) No Bartlett's statistic (corrected) P value Significantly different standard deviations? (P <.5) ANOVA table SS DF MS F (DFn, DFd) P value Treatment (between columns) F (, ) = 5.59 P <. Residual (within columns) Total.567 Number of treatments (columns) 5 Number of values (total) 5 Number of families Number of comparisons per family Alpha.5 Tukey's multiple comparisons test Mean Diff. 95% CI of diff. Significant? Summary vs. shtert-a.5 -. to.8 No ns

23 vs. shtert-b.5.7 to.995 Yes * vs. shtert-a +TERT to -.7 Yes * vs. shtert-b +TERT to.966 No ns shtert-a vs. shtert-b to.5 No ns shtert-a vs. shtert-a +TERT to -.9 Yes * shtert-a vs. shtert-b +TERT to Yes * shtert-b vs. shtert-a +TERT to Yes * shtert-b vs. shtert-b +TERT to -.57 Yes * shtert-a +TERT vs. shtert-b +TERT to.99 No ns LDHA F.76 P value <. Are differences among means statistically significant? (P <.5) Yes R square.897 F (DFn, DFd).79 (, ) P value.7 ns Significantly different standard deviations? (P <.5) No Bartlett's statistic (corrected) P value Significantly different standard deviations? (P <.5) ANOVA table SS DF MS F (DFn, DFd) P value Treatment (between columns).9.9 F (, ) =.76 P <. Residual (within columns).8.8 Total. Number of treatments (columns) 5 Number of values (total) 5 Number of families Number of comparisons per family Alpha.5 Tukey's multiple comparisons test Mean Diff. 95% CI of diff. Significant? Summary vs. shtert-a to.77 Yes * vs. shtert-b to.797 Yes vs. shtert-a +TERT to. No ns vs. shtert-b +TERT to.8 No ns shtert-a vs. shtert-b to.99 No ns shtert-a vs. shtert-a +TERT to -.7 Yes * shtert-a vs. shtert-b +TERT to -.67 Yes shtert-b vs. shtert-a +TERT to -.59 Yes * shtert-b vs. shtert-b +TERT to -.6 Yes shtert-a +TERT vs. shtert-b +TERT. -. to.56 No ns PKM F.66 P value <. Are differences among means statistically significant? (P <.5) Yes R square.8965 F (DFn, DFd).7 (, ) P value.9 ns Significantly different standard deviations? (P <.5) No Bartlett's statistic (corrected) P value Significantly different standard deviations? (P <.5) ANOVA table SS DF MS F (DFn, DFd) P value Treatment (between columns).. F (, ) =.66 P <. Residual (within columns) Total.9 Number of treatments (columns) 5

24 Number of values (total) 5 Number of families Number of comparisons per family Alpha.5 Tukey's multiple comparisons test Mean Diff. 95% CI of diff. Significant? Summary vs. shtert-a to.6695 Yes * vs. shtert-b to.7 Yes * vs. shtert-a +TERT to.8 No ns vs. shtert-b +TERT to.97 No ns shtert-a vs. shtert-b to.679 No ns shtert-a vs. shtert-a +TERT to -. Yes * shtert-a vs. shtert-b +TERT to Yes * shtert-b vs. shtert-a +TERT to -.6 Yes * shtert-b vs. shtert-b +TERT to -. Yes * shtert-a +TERT vs. shtert-b +TERT to.7 No ns EXOSC F 5.9 P value <. Are differences among means statistically significant? (P <.5) Yes R square.98 F (DFn, DFd).96 (, ) P value.8 ns Significantly different standard deviations? (P <.5) No Bartlett's statistic (corrected) P value Significantly different standard deviations? (P <.5) ANOVA table SS DF MS F (DFn, DFd) P value Treatment (between columns) F (, ) = 5.9 P <. Residual (within columns).6.6 Total.66 Number of treatments (columns) 5 Number of values (total) 5 Number of families Number of comparisons per family Alpha.5 Tukey's multiple comparisons test Mean Diff. 95% CI of diff. Significant? Summary vs. shtert-a to.88 Yes * vs. shtert-b.8.7 to.5 Yes vs. shtert-a +TERT to.96 No ns vs. shtert-b +TERT to. No ns shtert-a vs. shtert-b to.56 No ns shtert-a vs. shtert-a +TERT to Yes shtert-a vs. shtert-b +TERT to Yes * shtert-b vs. shtert-a +TERT to -.55 Yes shtert-b vs. shtert-b +TERT to -.5 Yes shtert-a +TERT vs. shtert-b +TERT to.58 No ns MCM6 F 7.8 P value.5 Are differences among means statistically significant? (P <.5) Yes R square.75 F (DFn, DFd).98 (, ) P value.699 ns Significantly different standard deviations? (P <.5) No Bartlett's statistic (corrected) P value

25 Significantly different standard deviations? (P <.5) ANOVA table SS DF MS F (DFn, DFd) P value Treatment (between columns) F (, ) = 7.8 P =.5 Residual (within columns).9.9 Total.855 Number of treatments (columns) 5 Number of values (total) 5 Number of families Number of comparisons per family Alpha.5 Tukey's multiple comparisons test Mean Diff. 95% CI of diff. Significant? Summary vs. shtert-a to.6 No ns vs. shtert-b..7 to.88 Yes * vs. shtert-a +TERT to.99 No ns vs. shtert-b +TERT to.678 No ns shtert-a vs. shtert-b to.595 No ns shtert-a vs. shtert-a +TERT to.67 No ns shtert-a vs. shtert-b +TERT to.57 No ns shtert-b vs. shtert-a +TERT to -.95 Yes shtert-b vs. shtert-b +TERT to -.5 Yes * shtert-a +TERT vs. shtert-b +TERT to.559 No ns MYBBPA F.5 P value.9 * Are differences among means statistically significant? (P <.5) Yes R square.85 F (DFn, DFd).8 (, ) P value. ns Significantly different standard deviations? (P <.5) No Bartlett's statistic (corrected) P value Significantly different standard deviations? (P <.5) ANOVA table SS DF MS F (DFn, DFd) P value Treatment (between columns).969. F (, ) =.5 P =.9 Residual (within columns).6.6 Total. Number of treatments (columns) 5 Number of values (total) 5 Number of families Number of comparisons per family Alpha.5 Tukey's multiple comparisons test Mean Diff. 95% CI of diff. Significant? Summary vs. shtert-a to.78 No ns vs. shtert-b..67 to.795 Yes * vs. shtert-a +TERT to.9 No ns vs. shtert-b +TERT to.9 No ns shtert-a vs. shtert-b to.89 No ns shtert-a vs. shtert-a +TERT to -.5 Yes shtert-a vs. shtert-b +TERT to -.7 Yes shtert-b vs. shtert-a +TERT to -.98 Yes shtert-b vs. shtert-b +TERT to -.6 Yes shtert-a +TERT vs. shtert-b +TERT to.679 No ns

26 Sta$s$cs for Figure A HK F 8.65 P value <. Are differences among means statisticayes R square.7 F (DFn, DFd).5 (, 5) P value.667 ns Significantly different standard deviatio No Bartlett's statistic (corrected).95 P value. ns Significantly different standard deviatio No ANOVA table SS DF MS F (DFn, DFdP value Treatment (between columns).. F (, 5) = P <. Residual (within columns) Total Number of treatments (columns) Number of values (total) 8 Number of families Number of comparisons per family Alpha.5 Tukey's multiple comparisons test Mean Diff. 95% CI of d Significant? Summary vs. shtert to. Yes A-B vs. shtert + MYC.79. to Yes * A-C shtert vs. shtert + MYC to -Yes * B-C Test details Mean Mean Mean Diff. SE of diff. n n q DF vs. shtert vs. shtert + MYC shtert vs. shtert + MYC PGK F 5.89 P value. * Are differences among means statisticayes R square.679 F (DFn, DFd) 5.5 (, 5) P value.58 * Significantly different standard deviatio Yes Bartlett's statistic (corrected) 9. P value.5 * Significantly different standard deviatio Yes

27 ANOVA table SS DF MS F (DFn, DFdP value Treatment (between columns) F (, 5) = P =. Residual (within columns) Total Number of treatments (columns) Number of values (total) 8 Number of families Number of comparisons per family Alpha.5 Tukey's multiple comparisons test Mean Diff. 95% CI of d Significant? Summary vs. shtert.9. to. Yes A-B vs. shtert + MYC to No ns A-C shtert vs. shtert + MYC to -Yes * B-C Test details Mean Mean Mean Diff. SE of diff. n n q vs. shtert vs. shtert + MYC shtert vs. shtert + MYC ALDOC F.8 P value <. Are differences among means statisticayes R square.879 F (DFn, DFd).86 (, 5) P value. ns Significantly different standard deviatio No Bartlett's statistic (corrected). P value.9 ns Significantly different standard deviatio No ANOVA table SS DF MS F (DFn, DFdP value Treatment (between columns) F (, 5) = P <. Residual (within columns) Total Number of treatments (columns) Number of values (total) 8 Number of families Number of comparisons per family Alpha.5 Tukey's multiple comparisons test Mean Diff. 95% CI of d Significant? Summary vs. shtert.96.7 to. Yes A-B vs. shtert + MYC to No ns A-C shtert vs. shtert + MYC to -Yes B-C Test details Mean Mean Mean Diff. SE of diff. n n

28 vs. shtert vs. shtert + MYC shtert vs. shtert + MYC PKM F.97 P value.9 * Are differences among means statisticayes R square.6 F (DFn, DFd).767 (, ) P value.857 ns Significantly different standard deviatio No Bartlett's statistic (corrected).5 P value.55 ns Significantly different standard deviatio No ANOVA table SS DF MS F (DFn, DFdP value Treatment (between columns).57.6 F (, ) = P =.9 Residual (within columns) Total. 6 Number of treatments (columns) Number of values (total) 7 Number of families Number of comparisons per family Alpha.5 Tukey's multiple comparisons test Mean Diff. 95% CI of d Significant? Summary vs. shtert to. Yes A-B vs. shtert + MYC to No ns A-C shtert vs. shtert + MYC to -Yes B-C Test details Mean Mean Mean Diff. SE of diff. n n q vs. shtert vs. shtert + MYC shtert vs. shtert + MYC LDHA F.9 P value.5 Are differences among means statisticayes R square.666 F (DFn, DFd).7 (, ) P value. Significantly different standard deviatio Yes

29 Bartlett's statistic (corrected) P value Significantly different standard deviations? (P <.5) ANOVA table SS DF MS F (DFn, DFdP value Treatment (between columns).7. F (, ) = P =.5 Residual (within columns)..999 Total.9 Number of treatments (columns) Number of values (total) Number of families Number of comparisons per family Alpha.5 Tukey's multiple comparisons test Mean Diff. 95% CI of d Significant? Summary vs. shtert.7.95 to Yes A-B vs. shtert + MYC to No ns A-C shtert vs. shtert + MYC to -Yes * B-C Test details Mean Mean Mean Diff. SE of diff. n n q DF vs. shtert vs. shtert + MYC shtert vs. shtert + MYC

30 Sta$s$cs for Figure S5G Cyclin D F 8.5 P value <. Are differences among means statistically significant? (P <.5) Yes R square.979 F (DFn, DFd) 7.5 (5, ) P value. * Significantly different standard deviations? (P <.5) Yes Bartlett's statistic (corrected) 9.7 P value <. Significantly different standard deviations? (P <.5) Yes ANOVA table SS DF MS F (DFn, DFdP value Treatment (between columns) F (5, ) = P <. Residual (within columns) Total. 5 Number of treatments (columns) 6 Number of values (total) 6 Number of families Number of comparisons per family 5 Alpha.5 Tukey's multiple comparisons test Mean Diff. 95% CI of d Significant? Summary Control vs. MYC to -.Yes Control vs. MYC MUT to No ns Control vs. TERT to -.Yes Control vs. TERT + MYC to -.Yes Control vs. TERT + MYC MUT to No ns MYC vs. MYC MUT..797 to. Yes MYC vs. TERT to -Yes MYC vs. TERT + MYC to -.Yes MYC vs. TERT + MYC MUT to. Yes MYC MUT vs. TERT to -.Yes MYC MUT vs. TERT + MYC to -.Yes MYC MUT vs. TERT + MYC MUT to No ns TERT vs. TERT + MYC to -Yes TERT vs. TERT + MYC MUT..999 to. Yes TERT + MYC vs. TERT + MYC MUT to. Yes PKM F 78.6 P value <. Are differences among means statistically significant? (P <.5) Yes R square.987 F (DFn, DFd).9 (5, ) P value.56 ns Significantly different standard deviations? (P <.5) No Bartlett's statistic (corrected) 9.57 P value.5

31 Significantly different standard deviations? (P <.5) Yes ANOVA table SS DF MS F (DFn, DFdP value Treatment (between columns). 5. F (5, ) = 7P <. Residual (within columns) Total.96 5 Number of treatments (columns) 6 Number of values (total) 6 Number of families Number of comparisons per family 5 Alpha.5 Tukey's multiple comparisons test Mean Diff. 95% CI of d Significant? Summary Control vs. MYC to -.Yes Control vs. MYC MUT to No ns Control vs. TERT to -.Yes Control vs. TERT + MYC to -.Yes Control vs. TERT + MYC MUT to No ns MYC vs. MYC MUT to. Yes MYC vs. TERT to No ns MYC vs. TERT + MYC to -Yes MYC vs. TERT + MYC MUT to. Yes MYC MUT vs. TERT to -.Yes MYC MUT vs. TERT + MYC to -.Yes MYC MUT vs. TERT + MYC MUT to No ns TERT vs. TERT + MYC to No ns TERT vs. TERT + MYC MUT..87 to. Yes TERT + MYC vs. TERT + MYC MUT.6.66 to.6 Yes GAPDH F 9.7 P value <. Are differences among means statistically significant? (P <.5) Yes R square.965 F (DFn, DFd) 6.8 (5, ) P value. * Significantly different standard deviations? (P <.5) Yes Bartlett's statistic (corrected) 5.7 P value.77 Significantly different standard deviations? (P <.5) Yes ANOVA table SS DF MS F (DFn, DFdP value Treatment (between columns) F (5, ) = P <. Residual (within columns)..7 Total Number of treatments (columns) 6 Number of values (total) 6 Number of families Number of comparisons per family 5 Alpha.5 Tukey's multiple comparisons test Mean Diff. 95% CI of d Significant? Summary Control vs. MYC to -.Yes

32 Control vs. MYC MUT to -Yes * Control vs. TERT to -.Yes Control vs. TERT + MYC to -.Yes Control vs. TERT + MYC MUT to -Yes MYC vs. MYC MUT to. Yes MYC vs. TERT to -Yes * MYC vs. TERT + MYC to -Yes MYC vs. TERT + MYC MUT to. Yes MYC MUT vs. TERT to -.Yes MYC MUT vs. TERT + MYC to -.Yes MYC MUT vs. TERT + MYC MUT to -Yes * TERT vs. TERT + MYC to -Yes TERT vs. TERT + MYC MUT.7.5 to. Yes TERT + MYC vs. TERT + MYC MUT.9.75 to. Yes

33 Sta$s$cs for Figure S6C Cyclin D F 7.7 P value <. Are differences among means statistically significant? (P Yes R square.95 F (DFn, DFd).8 (5, 8) P value.6 ns Significantly different standard deviations? (P <.5) No Bartlett's statistic (corrected).9 P value.95 * Significantly different standard deviations? (P <.5) Yes ANOVA table SS DF MS F (DFn, DFdP value Treatment (between columns) F (5, 8) = 7P <. Residual (within columns) Total 9.69 Number of treatments (columns) 6 Number of values (total) Number of families Number of comparisons per family 5 Alpha.5 Tukey's multiple comparisons test Mean Diff. 95% CI of d Significant? Summary + Vector vs. + MYC to No ns + Vector vs. + MYC T58A to -.Yes + Vector vs. shtert + Vector..67 to Yes * + Vector vs. shtert + MYC to No ns + Vector vs. shtert + MYC T58A to -.Yes + MYC vs. + MYC T58A to -Yes * + MYC vs. shtert + Vector.7.65 to. Yes + MYC vs. shtert + MYC to No ns + MYC vs. shtert + MYC T58A to -.Yes + MYC T58A vs. shtert + Vector to. Yes + MYC T58A vs. shtert + MYC.577. to. Yes + MYC T58A vs. shtert + MYC T58A to -.Yes shtert + Vector vs. shtert + MYC to -Yes * shtert + Vector vs. shtert + MYC T58A to -.Yes shtert + MYC vs. shtert + MYC T58A to -.Yes eifa F.5 P value <. Are differences among means statistically significant? (P Yes R square.975 F (DFn, DFd).85 (5, 8) P value.7 * Significantly different standard deviations? (P <.5) Yes Bartlett's statistic (corrected) 5. P value.99 Significantly different standard deviations? (P <.5) Yes

34 ANOVA table SS DF MS F (DFn, DFdP value Treatment (between columns) F (5, 8) = P <. Residual (within columns) Total 9. Number of treatments (columns) 6 Number of values (total) Number of families Number of comparisons per family 5 Alpha.5 Tukey's multiple comparisons test Mean Diff. 95% CI of d Significant? Summary + Vector vs. + MYC to No ns + Vector vs. + MYC T58A to -.Yes + Vector vs. shtert + Vector to No ns + Vector vs. shtert + MYC to No ns + Vector vs. shtert + MYC T58A to -.Yes + MYC vs. + MYC T58A to -.Yes + MYC vs. shtert + Vector to. Yes + MYC vs. shtert + MYC to No ns + MYC vs. shtert + MYC T58A to -.Yes + MYC T58A vs. shtert + Vector.56. to. Yes + MYC T58A vs. shtert + MYC to. Yes + MYC T58A vs. shtert + MYC T58A to No ns shtert + Vector vs. shtert + MYC to No ns shtert + Vector vs. shtert + MYC T58A to -.Yes shtert + MYC vs. shtert + MYC T58A to -.Yes NCL F 5.89 P value <. Are differences among means statistically significant? (P Yes R square.988 F (DFn, DFd).76 (5, 8) P value.7 * Significantly different standard deviations? (P <.5) Yes Bartlett's statistic (corrected) 9.97 P value. Significantly different standard deviations? (P <.5) Yes ANOVA table SS DF MS F (DFn, DFdP value Treatment (between columns) F (5, 8) = P <. Residual (within columns) Total.8 Number of treatments (columns) 6 Number of values (total) Number of families Number of comparisons per family 5 Alpha.5 Tukey's multiple comparisons test Mean Diff. 95% CI of d Significant? Summary + Vector vs. + MYC to.no ns + Vector vs. + MYC T58A to -.Yes + Vector vs. shtert + Vector to No ns + Vector vs. shtert + MYC to No ns + Vector vs. shtert + MYC T58A to -.Yes + MYC vs. + MYC T58A to -.Yes

35 + MYC vs. shtert + Vector to. Yes * + MYC vs. shtert + MYC to No ns + MYC vs. shtert + MYC T58A to -.Yes * + MYC T58A vs. shtert + Vector to. Yes + MYC T58A vs. shtert + MYC.8.69 to. Yes + MYC T58A vs. shtert + MYC T58A to No ns shtert + Vector vs. shtert + MYC to -.Yes * shtert + Vector vs. shtert + MYC T58A to -.Yes shtert + MYC vs. shtert + MYC T58A to -.Yes

36 A Wild-type EµMYC B shtert-a shtert-b. Figure S C Mouse ^ * TERT MYC ACTIN Relative Viability Control Relative Viability.8.6. shtert-a shtert-b. D Wildtype B EuMYC EµMYC B B Cells B cells Cells EµMYC lymphoma cells E sh-control sh-terc sh-tert-a sh-control sh-terc TERT Relative Terc Expression TERT ACTIN shterc

37 Figure S A EµMYC TERT +/+ EµMYC TERT -/- EµMYC TERT +/+ EµMYC TERT -/- B EµMYC TERT +/+ B cells EµMYC TERT -/- B cells DAPI TRF γhax

38 SENP NM_567 LDHB NM_797 LDHB NM_ NRD NM_7 SPRYD NM_7 LDHA NM_59 LDHA NM_655 LDHA NM_656 LDHA NM_5566 PFKFB NM_566 EIFA NM_5 EIFA NM_6 WDR NM_5 LDHA NM_65 GLS NM_8796 GLS NM_8797 GLS NM_8798 GLS NM_67 GPSM NM_568 GPSM NM_569 GPSM NM_ DUSL NM_669 DUSL NM_75 YARS NM_68 MRPL NM_78 POLE NM_7855 POLE NM_7 DNLZ NM_889 YDJC NM_796 EIF5A NM_76 EIF5A NM_76 EIF5A NM_76 EIF5A NM_97 SPBP NM_56 SPBP NM_75 C9orf NM_7869 C9orf NM_786 C9orf NM_5786 CARD9 NM_58 CARD9 NM_58 CCDC6 NM_56 GPS NM_89 Gamt NM_55 Atraid NM_7855 Atraid NM_7 Pemt NM_9 Pemt NM_9 Rfc NM_58 Bodl NM_8 Shtc NM_9 Dazap NM_6 Dazap NM_65 Dazap NM_88 Cad NM_895 Cad NM_895 Cad NM_55 Shcbpl NM_6 Gar NM_6578 Tbrg NM_57 Tbrg NM_ Prrl NM_9 Dnajb NM_787 Atp5g NM_668 Jag NM_588 Ppia NM_897 Fubp NM_577 A Input shterc shtert Input shterc shtert B WT Input WT Tert / Figure S WT Input WT Tert / Input shterc shtert Input shterc shtert WT Input WT Tert / WT Input WT Tert / Input shterc shtert Input shterc shtert WT Input WT Tert / WT Input WT Tert / Input shterc shtert Input shterc shtert WT Input WT Tert / WT Input WT Tert / Input shterc shtert Input shterc shtert WT Input WT Tert / WT Input WT Tert / Input shterc shtert Input shterc shtert WT Input WT Tert / WT Input WT Tert / Input shterc shtert Input shterc shtert WT Input WT Tert / WT Input WT Tert / C D % Input shtert shtert shtert shtert + TERT WT shtert + TERT DN Nucleolin Cyclin D eifα # % Input H # shtert shtert shtert shtert + TERT WT shtert + TERT DN

39 E % Input MYC ChIP: Eu-MYC;TERT+/+ MYC ChIP: Eu-MYC;TERT-/- Pol ChIP: Eu-MYC;TERT+/+ Pol ChIP: Eu-MYC;TERT-/- * ^ ^ * ^ * * * * % Input F ^ NF-Y ChIP ^ ^ Figure S shtert ZNF77 JMJDB NLK RUVBL CONTROL ^ G TERT 5s rrna 5.8s rrna 5s rrna 8s rrna 8s rrna GAPDH ENO LDHA PKM EXOSC GART CCT7 MCM5 I Relative Expression..... TERT+/+; EuMYC ^ TERT+/-; EuMYC TERT-/-; EuMYC Relative Expression Cyclin D eifα Nucleolin shterc shtert shtert shtert shtert + TERT WT shtert + TERT DN Relative Expression J Relative Expression H * 5s rrna # shterc shtert shtert shtert shtert + TERT WT shtert + TERT DN ^ shtert-a shtert-b shtert-a +TERT * * shtert-b +TERT * * * * *

40 Figure S A.. High MYC Rela%ve TERT Expression * * *.. shtert shtert shtert B.. Low MYC Rela%ve TERT Expression * * *.. shtert shtert shtert

41 A Relative TERT Expression shtert shtert MYC ACTIN Figure S5 B MIHA INPUT IP: IgG IP: MYC TERT MYC MAX ACTIN C Input shtert sh Terc IP: IgG shtert sh Terc IP: MYC shtert sh Terc TERT MYC MAX ACTIN

42 D NTD c-myc CTD MBI MBII MBIII AR NLS B HLH LZ Figure S E Input IP: FLAG pbobi Flag vector pcmv Flag MYC CβF-MYC - * CβF-MYC 9-5 CβF-MYC -8 CβF-MYC - CβF-MYC -8 CβF-MYC 8-8 TERT FLAG TERT FLAG F TT Control TT+ GST TT+ MYC (-8) TT+ MYC (98-9) Control+ GST Control+ MYC (-8) COntrol+ MYC (98-9) 5 6 IP: GST IB: TERT Marker GST pgext-myc (-8) pgexx-myc (98-9) Marker 5 Comassie Gel G Relative Expression ^ ^ Vector MYC MYC Δ- TERT MYC + TERT MYC Δ- + TERT * Cyclin D GAPDH PKM ^ ^ H Input IP: FLAG Flag vector WT TERT DN TERT -6 TERT 6-7 TERT -95 TERT 5-95 TERT MYC FLAG FLAG ACTIN 5 6 7

43 A shtert shtert B + Flag-MYC T58A shtert + Flag-MYC T58A Figure S6 CHX (min) MYC CHX(min) 6 6 FLAG (MYC T58A) TERT TERT ACTIN 5 6 Relative Expression C Vector + MYC + MYC T58A shtert + Vector shtert +MYC shtert + MYC T58A ^. ^.5 ^ ^ ^ ^ D HA-Ub shtert Flag-MYC IP: FLAG HA. eifα Nucleolin Cyclin D HA E IP: HIS Input Input shtert shtert + His-ubiquitin + Flag MYC Δ- TERT TERT Flag ACTIN FLAG FLAG ACTIN