Isolate Sexual Idiomorph Species
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1 SUPLEMENTARY TABLE 1. Isolate identification, sexual idiomorph and species of each isolate used for MAT locus distribution in Paracoccidioides species. Isolate Sexual Idiomorph Species Pb01 MAT1-1 P. lutzii 3171 MAT1-2 P. lutzii EE MAT1-2 P. lutzii RAJ-2 MAT1-2 P. lutzii Pb3 MAT1-2 P. brasiliensis PS2 Pb73 MAT1-1 P. brasiliensis PS3 Pb18 MAT1-1/MAT1-2 P. brasiliensis S1 Pb2 MAT1-1 P. brasiliensis PS2 Pb4 MAT1-1 P. brasiliensis PS2 192 MAT1-1 N.I. 113 MAT1-1 N.I. Higuita/P196 MAT1-2 P. brasiliensis PS3 Beto MAT1-1 N.I. 135 MAT1-2 P. brasiliensis S1 300 MAT1-1/MAT1-2 P. brasiliensis S MAT1-1/MAT1-2 P. lutzii 6810 MAT1-1 P. lutzii MND MAT1-1 P. lutzii JMS MAT1-2 P. lutzii 728 MAT1-1/MAT1-2 N.I MAT1-1/MAT1-2 N.I. Edi MAT1-1 N.I 68 MAT1-1 N.I. SN MAT1-1 N.I. JT-5 MAT1-1 N.I. Givaldo MAT1-1 N.I 1684 MAT1-1 P. brasiliensis S1 PbBT MAT1-1 P. brasiliensis S1 662 MAT1-2 N.I. JHS MAT1-2 P. lutzii 769 MAT1-1 N.I.* 133 MAT1-1/MAT1-2 N.I.* 397 MAT1-1 P. lutzii 307 MAT1-1/MAT1-2 P. barsiliensis S MAT1-1 P. brasiliensis S1 305 MAT1-1 P. brasiliensis S1 Pingüim MAT1-1 N.I. 351 MAT1-1/MAT1-2 P. lutzii 5979 MAT1-1/MAT1-2 P. lutzii 698 MAT1-2 N.I. JAL MAT1-1 P. lutzii 52 MAT1-1 N.I. 53 MAT1-2 N.I. TATU MAT1-2 N.I. 262 MAT1-2 N.I 1578 MAT1-1 P. lutzii 8434 MAT1-1 P. lutzii
2 5979 MAT1-1 P. lutzii 6184 MAT1-1 N.I. 16 MAT1-2 N.I. D03 MAT1-2 N.I. D01 MAT1-2 N.I. 265 MAT1-2 N.I. EPM101 MAT1-1 N.I. EPM77 MAT1-2 N.I. BT84 MAT1-1 P. brasiliensis PS2 BT60 MAT1-1 P. brasiliensis S1 S1 Baço MAT1-1 N.I. T9B1 MAT1-1 P. brasiliensis S1 T8LN2 MAT1-1 N.I. T7B6 MAT1-1 P. brasiliensis S1 T3B6 MAT1-1 P. brasiliensis S1 T1F1 MAT1-1 P. brasiliensis S1 EPM75 MAT1-1 N.I. D11 MAT1-1 N.I. D10 MAT1-2 N.I. D07 MAT1-2 N.I. D06 MAT1-2 N.I. MFC MAT1-1 P. lutzii 309 MAT1-1 P. brasiliensis S1 P934 MAT1-1 N.I MAT1-1 N.I. 66 MAT1-1 P. lutzii 8934 MAT1-1 P.lutzii T4B17 MAT1-1 N.I. BT85 MAT1-1/MAT1-2 N.I. T10B1 MAT1-2 P. brasiliensis PS2 T13LN2 MAT1-1 N.I. ILSL57 MAT1-2 N.I. CÃO MAT1-1 P. brasiliensis PS MAT1-1 N.I. D02 MAT1-1 N.I. D04 MAT1-1 N.I. D09 MAT1-1 N.I. D05 MAT1-2 N.I. EPM04 MAT1-1/MAT1-2 N.I. EPM85 MAT1-2 N.I. EPM83 MAT1-2 N.I. Pb21 MAT1-2 N.I. ED01 MAT1-1 P. lutzii PLRO1 MAT1-2 P. lutzii Pb9 MAT1-1 P. brasiliensis S1 Pb339 MAT1-1 P. brasiliensis S1 T15LN1 MAT1-2 P. brasiliensis S1 JT-4 MAT1-1 N.I. T8B1 MAT1-2 P. brasiliensis S MAT1-1/MAT1-2 N.I.
3 262 MAT1-2 N.I. N.I. Paracoccidioides species not identified neither by molecular or morphological data. * 769 and 133 isolates share alleles with both P. brasiliensis and P. lutzii species and could not be placed in any species by genealogical concordance method
4 Supplementary Table 2. Primers desigend for MAT locus amplification and Real time RT-PCR for sex related genes in Paracoccidioides strains. Primers designed to amplify the MAT locus Locus Primer Direction Sequence α-box (MAT1-1) MAT1-1F Foward 5' GCC AGG ATG ATA GGA AAC GAA GTC 3' MAT1-1R Reverse 5' GGG AAA GAT CCG GGA ATA CAG TAG 3' HMG (MAT1-2) MAT1-2F Foward 5' GAA GGC TTT CGA GGA GCA TAT TGA TAG 3' MAT1-2R Reverse 5' GAG TGC TAT GAA CCA TCT GAA GTC TAC 3' Primer designed for Real time RT-PCR Locus Primer Direction Sequence α-box (MAT1-1) RETIalphaF Foward 5' CCC GGA TCT TTC CCA AGA AG 3' RETIalphaR Reverse 5' GGA TTG CCC ACT TTG CTT TG 3' HMG (MAT1-2) RETIHMGF Foward 5' CAA ACG CCA ATT TCC TCT CAT C 3' RETIHMGF Reverse 5' AGC TGC GGA CTC GGT ATA AGA G 3' HMG (MAT1-2)* HMGlutziiF Foward 5' CGT TCA AAG ACA ACG TGA ATG G 3' HMGlutziiR Reverse 5' TCA TCC GTG GGC CAG ATA AG 3' ste3 ste3f Foward 5' TTT CCT GCA TCC TGC CCT AT 3' ste3r Reverse 5' TCA TCC GTG GGC CAG ATA AG 3' ste2 ste2f Foward 5' GGT TTG GTG CTA TGC AGG TTA TG 3' ste2r Reverse 5' GGA GTG GGT ATT GGA GAA TTG AGA 3' fus3 fus3f Foward 5' AGG ACC TTT CCG ACG ACC AT 3' fus3r Reverse 5' AAC GTT CGC AGA GTG CAT AGC 3' ste12 ste12f Foward 5' GGA TTT GGG AAG TAA CCA CTC TAC C 3' ste12r Reverse 5' CCC AGC ACC AGT CAC AGT AAA G3' * Primer designed for P. lutzii strains
5 SUPLEMENTARY TABLE 3 - Pairwise mating experiements between and within P. lutzii and P. brasiliensis species. Isolates used and young ascocarp formation (frequency of coiled hyphae by slide analyzed) are indicated in each row. Isolates Young ascocarp formation* P. lutzii mating assays JAL X EE X EE X EE + ED01 X PLRO1 ++ Pb01 X EE - P. brasiliensis mating assays T1F1 X T15LN1 +++ T9B1 X T15LN1 +++ Pb2 X Pb3 - Pb4 X Pb3 - Pb18 X T9B1 - P. lutzii X P. brasiliensis mating assays T1F1 X EE - JAL X T15LN1 - * +++, more than 25 coiled hyphae per slide; ++, coiled hyphae per slide; + up-to 10 coiled hyphae per slide; - no coil observed;
6 SUPPLEMENTARY TABLE 4: Putative sex related genes shown by a BLAST searches on the dimorphic fungal genomes The table content shows the protein name, function in S. cerevisiae and the access number for all organisms identified gene function P. brasiliensis P. lutzii H. capsulatum B. dermatitidis C. immitis C. posadasi Pheromone receptors ste2 Pheromone receptor factor α PABG_ PADG_ PAAG_ BDBG_ BDCG_ CPAG_ CPSG_ ste3 gba1 ste4 Pheromone receptor factor a GTP-binding alpha subunit of the heterotrimeric G protein that couples to pheromone receptors; G protein beta subunit, forms a dimer with Ste18p to activate the mating signaling pathway, forms a heterotrimer with Gpa1p and Ste18p to dampen PADG_ PABG_ PADG_ PABG_ PAAG_ HCEG_ HCDG_ HCBG_ HCAG_ HCEG_ HCBG_ HCDG_ HCAG_ BDCG_ BDBG_ Components of a pheromone-response pathway involved in a mating process PAAG_ HCEG_ BDBG_ HCDG_ BDCG_ HCAG_ PADG_ PABG_ PAAG_ HCBG_ HCEG_ HCDG_ HCAG_ BDBG_ BDCG_ CIRG_ CISG_ CIMG_ CIHG_ CIRG_ CISG_ CIMG_ CIHG_ CIRG_ CISG_ CIMG_ CIHG_ CIRG_ CISG_ CIMG_ CIHG_ CPAG_ CPSG_ CPAG_ CPSG_ CPAG_ CPSG_
7 ste18 ste20 ste11 signaling G protein gamma subunit, forms a dimer with Ste4p to activate the mating signaling pathway, forms a heterotrimer with Gpa1p and Ste4p to dampen signaling Signal transducing kinase of the PAK (p21-activated kinase) family, involved in pheromone response and pseudohyphal/invasiv e growth pathways, activated by Cdc42p; binds Ste4p at a GBB motif present in noncatalytic domains of PAK kinases Serine/threonine protein kinase required for cell-typespecific transcription and signal transduction in yeast. PADG_ PABG_ PABG_ PADG_ PADG_ PABG_ PAAG_ PAAG_ PAAG_ HCBG_ HCDG_ HCAG_ HCEG_ HCBG_ HCEG_ HCAG_ HCEG_ HCAG_ HCBG_ BDBG_ BDCG_ BDCG_ BDBG_ BDBG_ BDCG_ CIRG_ CISG_ CIMG_ CIHG_ CISG_ CIRG_ CIMG_ CIHG_ CIRG_ CIMG_ CIHG_ CISG_ CPAG_ CPSG_ CPAG_ CPSG_ CPAG_ CPSG_
8 ste7 fus3 Signal transducing MEK kinase involved in pheromoneresponse and pseudohyphal/invasiv e growth pathways, where it phosphorylates Ste7p, and the high osmolarity response pathway, via phosphorylation of Pbs2p; regulated by Ste20p and Ste50p Signal transducing MAP kinase kinase involved in pheromone response, where it phosphorylates Fus3p, and in the pseudohyphal/invasiv e growth pathway, through phosphorylation of Kss1p Mitogen-activated serine/threonine protein kinase involved in mating; PADG_ PABG_ PADG_ PABG_ PAAG_ : PAAG_ HCAG_ HCEG_ HCBG_ HCBG_ HCEG_ HCAG_ BDBG_ BDCG_ BDBG_ BDCG_ CIHG_ CIRG_ CISG_ CIMG_ CIRG_ CISG_ CIMG_ CIHG_ CPAG_ CPSG_ CPAG_ CPSG_
9 phosphoactivated by Ste7p; substrates include Ste12p, Far1p, Bni1p, Sst2p ste12 PABG_ PADG_ PAAG_ HCBG_ HCDG_ HCAG_ BDBG_ BDCG_ CISG_ CIMG_ CIHG_ CIRG_ CPAG_ CPSG_ ram1 ram2 Beta subunit of the CAAX farnesyltransferase (FTase) that prenylates the a- factor mating pheromone and Ras proteins; required for the membrane localization of Ras proteins and a-factor Alpha subunit of both the farnesyltransferase and type I geranylgeranyltransfe rase that catalyze prenylation of proteins containing a CAAX consensus Pheromone-processing enzymes involved in a mating process PADG_ PAAG_ HCDG_ BDBG_ PABG_ HCBG_ BDCG_ HCEG_ HCAG_ PADG_ PABG_ PAAG_ HCDG_ HCEG_ HCBG_ HCAG_ BDBG_ BDCG_ CIHG_ CIRG_ CIMG_ CISG_ CIRG_ CISG_ CIMG_ CIHG_ CPAG_ CPSG_ CPAG_ CPSG_
10 rce1p ste24 ste14 motif; essential protein required for membrane localization of Ras proteins and a-factor Type II CAAX prenyl protease involved in the proteolysis and maturation of Ras and the a-factor mating pheromone Highly conserved zinc metalloprotease that functions in two steps of a-factor maturation, C- terminal CAAX proteolysis and the first step of N- terminal proteolytic processing Farnesyl cysteinecarboxyl methyltransferase, mediates the carboxyl methylation step during C-terminal CAAX motif processing of a-factor and RAS proteins in PABG_ PADG_ PABG_ PADG_ PADG_ PABG_ PAAG_ PAAG_ PAAG_ HCBG_ HCEG_ HCAG_ HCDG_ HCEG_ HCDG_ HCBG_ HCAG_ HCEG_ HCDG_ HCBG_ BDBG_ BDCG_ BDBG_ BDCG_ BDCG_ BDBG_ CIRG_ CIMG_ CISG_ CIRG_ CIMG_ CIHG_ CISG_ CIRG_ CISG_ CIMG_ CIHG_ CPAG_ CPSG_ CPAG_ CPSG_ CPAG_ CPSG_
11 ste6 kex1 kex2 the endoplasmic reticulum, localizes to the ER membrane Plasma membrane ATP-binding cassette (ABC) transporter required for the export of a-factor, catalyzes ATP hydrolysis coupled to a-factor transport; contains 12 transmembrane domains and two ATP binding domains; expressed only in MATa cells Protease involved in the processing of killer toxin and alpha factor precursor; cleaves Lys and Arg residues from the C- terminus of peptides and proteins Endoprotease α-factor processing PADG_ PABG_ PADG_ PABG_ PADG_ PABG_ PAAG_ PAAG_ PAAG_ HCBG_ HCEG_ HCAG_ HCDG_ HCEG_ HCAG_ HCDG_ HCBG_ HCBG_ HCAG_ HCEG_ HCDG_ BDBG_ BDCG_ BDBG_ BDCG_ BDCG_ BDBG_ CIRG_ CIMG_ CIHG_ CISG_ CIRG_ CIMG_ CIHG_ CIRG_ CIMG_ CIHG_ CISG_ CPAG_ CPSG_ CPAG_ CPSG_ CPAG_ CPSG_
12 ste13 Dipeptidyl aminopeptidase, Golgi integral membrane protein that cleaves on the carboxyl side of repeating -X-Alasequences, required for maturation of alpha factor, transcription is induced by a-factor PABG_ PADG_ PAAG_ HCBG_ HCEG_ HCDG_ HCAG_ BDBG_ BDCG_ CIMG_ CIRG_ CIHG_ CISG_ CPSG_ CPAG_
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