ion between daughters. pta2D shows strong genetic interactions with Cdc42 regulators and with key polarity factors Tea1, Tea4 and For3. Our results demonstrate a role for pta2 in PP2A dependent regulation of cell polarity and cell growth. Results and Discussion Identification of proteins associated with PP2A subunit paa1 In order to identify new regulators and/or substrates of the fission yeast PP2A complex we used co-immunoprecipitation followed by mass spectrometry analysis. The structural subunit of PP2A complex Paa1 was tagged at the C-terminus with the PK epitope at the endogenous chromosomal locus and was expressed under the control of its native promoter. The resulting protein fusion was functional as judged by the absence of any observable phenotype. Protein extracts from exponentially growing Paa1-PK expressing cells and untagged wild type cells as a control were subjected to immunoprecipitation with anti-PK tag antibody. Precipitated proteins were resolved by SDS polyacrylamide gel electrophoresis and detected using silver staining. The immunoprecipitation experiment was repeated four times and consistently observed bands BS-181 site corresponding to Paa1-PK associated proteins were excised from the gel and identified using tandem mass spectrometry. Among Paa1 associated proteins we found the two catalytic subunits of PP2A, 2 PP2A Role in S. pombe Morphogenesis Strain RD 312 RD 313 RD 1192 RD 1193 RD 1439 RD 821 RD 1349 RD 1292 RD 1298 RD 1306 RD 1304 RD 1139 RD 1176 RD 1178 RD 2064 RD 1346 RD 1266 RD 1262 RD 1294 RD 264 RD 1099 RD 1177 RD 1459 RD 1494 RD 1505 Genotype h+ ade6-M210 ura4-D18 leu1-32 h2 ade6-M216 ura4-D18 leu1-32 h+ pta2::ura4+ade6-M216 ura4-D18 leu1-32 h2 pta2::ura4+ade6-M216 ura4-D18 leu1-32 h+ paa1:pK ade6- leu1-32 h2 pta2:GFP:kanMX6 ade6-M216 ura4-D18 leu1-32 h pta2::ura4+tea1:GFP:kanMX6 ade6-M216 ura4-D18 leu1-32 h pta2::ura4+for3:3GFP:ura4+ade6-M216 ura4-D18 leu1-32 h pta2::ura4+rga4:GFP:kanMX6 ura4-D18 leu1-32 h2 pta2::ura4+CRIB:GFP:ura4+ura4-D18 leu1-32 h2 tea1:GFP:kanMX6 
ade6-M216 ura4-D18 leu1-32 h2 for3:3GFP-ura4+ade6-M216 leu1-32 ura4-D18 h2 rga4:GFP:kanMX6 ura4-D18 leu1-32 h2 CRIB:GFP-ura4+ura4-D18 leu1-32 h2 pta2::ura4+gef1::ura4+ade6-M216 ura4-D18 leu1-32 h+ gef1::ura4+ade6-M216 ura4-D18 leu1-32 h pta2::ura4+tea4::kanMX6 ade6- ura4-D18 leu1-32 h pta2::ura4+tea1::ura4+ade6-M216 ura4-D18 h PubMed ID:http://www.ncbi.nlm.nih.gov/pubmed/22189475 pta2::ura4+rga4::ura4+ura4-D18 leu1-32 h+ tea1::ura4+ura4-D18 ade6-M216 h+ tea4::kanMX6 ade6- leu1-32 ura4-D18 h2 rga4::ura4+ura4-D18 lue1-32 h2 for3::kanMX6 ade6- ura4-D18 leu1-32 h+ HA:cdc42 ade6-M16 leu1-32 ura4-D18 h pta2::ura4+HA:cdc42 ade6- ura4-D18 leu1-32 Source Lab collection Lab collection This study This study This study This study This study This study This study This study Lab collection Martin SG. et al.,2005 Tatebe H. et al., 2008 Tatebe H. et al., 2008 This study Lab collection This study This study This study Mata J. et al., 1997 Martin SG. et al.,2005 Tatebe H et al., 2008 Lab collection Coll et all., 2005 This study doi:10.1371/journal.pone.0032823.t001 Ppa1 and Ppa2. We also found Par1, a regulatory B9 subunit of PP2A, and two genes not characterized in S. pombe, SPBP8B7.08c and SPAC1782.05. SPBP8B7.08c is a homologue of Ppm1, a leucine carboxyl methyltransferase that in other organisms is required for PP2A complex assembly and activity, while SPAC1782.05 is a homologue of the PP2A phosphatase activator . A number of other putative Paa1-PK interacting proteins, some of which may be substrates of the PP2A c
