Supplemental Fig. 1. Relative mrna Expression. Relative mrna Expression WT KO WT KO RT 4 0 C

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Supplemental Fig. 1 A 1.5 1..5 Hdac11 (ibat) n=4 n=4 n=4 n=4 n=4 n=4 n=4 n=4 WT KO WT KO WT KO WT KO RT 4 C RT 4 C Supplemental Figure 1. Hdac11 mrna is undetectable in KO adipose tissue. Quantitative PCR analysis of Hdac11 mrna expression in mouse ibat (A) and ingwat (B) tissues. Values represent means ±SEM. B 1.5 1..5 Hdac11 (ingwat)

Supplemental Fig. Plasma Concentration (pg/ml) 1 1 8 6 4 1 Epinephrine Norepinephrine n=4 WT n=4 n=4 n=4 KO WT KO Supplemental Figure. HDAC11 deficient mice have higher levels of circulating norepinephrine. Plasma catecholamine (epinephrine and norepinephrine) concentrations in 1 week-old male mice fed normal chow were determined by ELISA. Values for the graph represent means ±SEM; P<.5 vs. WT. Student s t-test was used for statistical analysis.

Supplemental Fig. 3 A Lenti-Control or Lenti-FLAG- HDAC11 (WT or H143A) B BRD4 (Co-IPed) Control WT H143A HEK93 Cells IP: FLAG IB: Endogenous BRD4 BRD4 (Input) FLAG-HD11 (Input) kda 5 15 Supplemental Figure 3. HDAC11 does not associate with BRD4. (A) Schematic representation of the HDAC11:BRD4 coimmunoprecipitation (IP) assay; IB, immunoblot. (B) Immunoblotting revealed the absence of BRD4 in HDAC11 immunoprecipitates. Total amounts of HDAC11 and endogenous BRD4 in homogenates were also determined (input).

Supplemental Fig. 4 A Lenti-Control or Lenti-FLAG- HDAC11 (H143A) HIB1B Brown Preadipocytes BAT Diff Medium (4 days) B 18 16 14 1 1 8 6 4 Ucp1 Diff: + + Ctrl H143A C 6 5 4 3 1 Pgc1α Diff: + + Ctrl H143A Supplementary Figure 4. Catalytically inactive HDAC11 fails to suppress thermogenic gene expression. (A) Schematic representation of the experiment. The H143A amino acid substitution renders HDAC11 catalytically inactive. (B-C) Quantitative PCR analysis of Ucp1 and Pgc1α mrna expression; n = 4 plates of cells/condition. Values for all graphs represent means ±SEM; P<.5 vs. Lenti-Control undifferentiated. Twoway ANOVA with Tukey s multiple comparisons test was employed.

Supplemental Fig. 5 A B Lenti-Control or Lenti-Myc- BRD (WT) or Lenti-Myc- BRD 1-6 -/+ Lenti-shHDAC11 HIB1B Brown Preadipocytes BAT Diff Medium (4 days) 15 1 5 Ucp1 Diff: + + + BRD WT: + + BRD 1-6: + shcontrol: + + + + shhdac11: + + # Supplementary Figure 5. BRD-mediated suppression of brown adipocyte differentiation requires association with HDAC11. (A) Schematic representation of the experiment. (B-C) Quantitative PCR analysis of Ucp1 and Pgc1α mrna expression; n = 4 plates of cells/condition. Values for all graphs represent means ±SEM; P<.5 vs. Lenti-Control undifferentiated; # P<.5 vs. Lenti-Control differentiated; P<.5 vs. Lenti-BRD WT differentiated. Two-way ANOVA with Tukey s multiple comparisons test was employed. C 5 4 3 1 Pgc1α Diff: + + + BRD WT: + + BRD 1-6: + shcontrol: + + + + shhdac11: + + #

Supplemental Fig. 6 A B Lenti-Control or Lenti-FLAG HDAC11 (WT or H143A) BAT Diff Medium (4 days) HIB1B Brown Preadipocytes IP Ac-H3K7 PCR Ucp1 Enhancer -57-463 Enrichment Relative to IgG IP 4 3 1 Chromatin Immunoprecipitation Diff: + + + Ctrl # WT H143A Supplementary Figure 6. HDAC11 blunts H3K7 acetylation of a Ucp1 enhancer element. (A) Schematic representation of the chromatin immunoprecipitation (ChIP) experiment. (B) ChIP-qPCR analysis showing relative enrichment of H3K7 acetylation at the Ucp1 enhancer. Values for the graph represent means ±SEM; P<.5 vs. Control undifferentiated; # P<.5 vs. control differentiated. Two-way ANOVA with Tukey s multiple comparisons test was employed.

Supplemental Tables Supplemental Table 1: Primers for qpcr Target 5 oligo 3 oligo Ucp1 CCGAAACTGTACAGCGGTCT CCGAGAGAGGCAGGTGTTTC Pgc1 TGTGTGCTGTGTGTCAGAGT ACCAGAGCAGCACACTCTATG 3-AR GGAAGCTTGCTTGATCCCCA GCCGTTGCTTGTCTTTCTGG Hdac11 TTACAACCGCCACATCTACC GACATTCCTCTCCACCTTCTC Brd AGCTGGGTTTGCCGGATTAT GGACAATATCATCGGTGGGCT Ucp1 enhancer (ChIP) AGCGTCACAGAGGGTCAGTC GTGAGGCTGATATCCCCAGA Supplemental Table : Primers for cloning Target 5 oligo 3 oligo BRD (1-6) CCATCGATCTGCAAAACGTGA CTCCCCACAATAAG CCGCTGAGTTATAAAGCAGCACT GCCACCCCCAC HDAC11 (H143A) GGGGGTGGCTTCCACGCCTG CTCCAGCGACCGT ACGGTCGCTGGAGCAGGCGTGG AAGCCACCCCC HDAC11 (1-3) CCATCGATCTACACACAACCC AGCTGTACCAGCAT GCTCTAGATCAGGTCACCATAAG GATGGGCACCCG HDAC11 (1-5) CCATCGATCTACACACAACCC AGCTGTACCAGCAT GCTCTAGATCACAGGTGCTCCTG GAGGGATTTCTT HDAC11 (1-) CCATCGATCTACACACAACCC AGCTGTACCAGCAT GCTCTAGATCAGATGTACACACG CTTGTCGTCCAT Supplemental Table 3: Primers for genotyping Target 5 oligo 3 oligo HDAC11 WT TGCTGCCTGTGAGCCACTGC AGAATGGCTGTCTCCCTAGG HDAC11 KO TGCTGCCTGTGAGCCACTGC CCTTGGAATAGCATCTCAGG Supplemental Table 4: Patient characteristics Brown Adipose Tissue-Deep Neck Patient ID Diagnosis Age Gender BMI (years) BAT-3 Multinodular goiter/ follicular adenoma 81 Female 37.7 BAT-9 Multinodular goiter 49 Male 34.4 BAT-13 Papillary thyroid carcinoma 7 Female 4. BAT-18 Parathyroid adenoma 56 Female 19. BAT-19 Benign follicular nodules 39 Female 6.9 BAT- Parathyroid adenoma 63 Female 4.8 1

White Adipose Tissue-Abdominal subcutaneous Patient ID Age Gender BMI (years) Ori-3 49 Female 3.9 Ori-33 34 Female 34.7 Ori-38 4 Female.3 Ori-54 51 Male 3. Ori-7 3 Male.4 Ori-76 9 Female 33.9 Ori-78 38 Female 37.9 Ori-79 7 Female 1.1 Ori-81 4 Female 1.1

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