E crossed the WT and chs1 query strains with an array plate containing all the genes of this chromosomal region. This experiment showed that chs1 combined with a deletion in any gene of this region, whether coding for an MSP or not, had a very negative S score. As shown in Fig 11C, plates showed a regional reduction of colony sizes for genes on Chr. II, whereas the crosses involving genes located on other Pinometostat web chromosomes (not boxed), showed no difference of colony sizes between the WT query plate and the chs1 query plate. In double mutants of chs1 combined with chs2-DAmP, fig1, fat1, cst26, or qdr3, we amplified by genomic PCR all genes and intergenic regions starting from CHS2 up to QDR3 and found that there were no rearrangements or deletions present apart from the intended single gene deletion. Interestingly, cst26 is one of the gene deletions sitting in the middle of the chromosomal region where deletions show negative S scores with chs1 (Fig 11A). As mentioned above, elo2 and elo3 also have very negative S scores in combination with cst26, similar to chs1 (Fig 11A). In this case however only very few genes immediately adjacent to cst26 show a negative S score with elo2 or elo3. The phenomenon of regionally concentrated negative interactions shown in Fig 11A is not an isolated phenomenon, since several such regions can easily be identified on a heat map of SPLOS Genetics | DOI:10.1371/journal.pgen.July 27,17 /Yeast E-MAP for Identification of Membrane Transporters Operating Lipid Flip FlopFig 11. Chs1-interacting cluster on Q-VD-OPh msds chromosome II. (A) correlation and selected interaction heat maps of a series of linked genes on the right arm of Chr. II, deletion of which causes significant negative interactions with chs1. Genes are in the same order as they are situated along the chromosome. (B) colonies present on MSP-E-MAP plates after the last selection of double mutants combining deletions in genes indicated on top of column and indicated to the left. As indicated, double mutant colonies are from crosses with chs1 used as query or being on an array. Tda5 was added as a neutral control, having an S score of 0.07 with chs1. (C) Shows plates from quadruplicate crosses of either a WT query (upper image) or chs1 query strain (lower image) with an array plate containing many genes from different chromosomes including all the genes from YBR020W to YBR078W on the right arm of Chr. II. All genes in this chromosomal region are boxed, in red if there is a size difference visible to the naked eye between the WT and chs1 plate, in yellow if there isn’t. Boxes are dotted for MSP genes that are part of the E-MAP set and are shown in panel A. doi:10.1371/journal.pgen.1006160.gscores where the genes are ordered according to their chromosomal location as shown in Fig 12. As the order in this matrix clusters each gene with the genes that sit next to it on the chromosome, all the irrelevant very negative interactions generated by proximity of two deletions on a same chromosome (< 100 kb) and hence marked with grey dots are clustering along the diagonal (Fig 12). Uniform interactions of all deletions in certain chromosomal regions withPLOS Genetics | DOI:10.1371/journal.pgen.July 27,18 /Yeast E-MAP for Identification of Membrane Transporters Operating Lipid Flip FlopFig 12. Interaction score heat map with genes ordered according to their chromosomal location. Standard names were replaced by the systematic names in the MSP-E-MAP interaction matrix (S3A Table) and.E crossed the WT and chs1 query strains with an array plate containing all the genes of this chromosomal region. This experiment showed that chs1 combined with a deletion in any gene of this region, whether coding for an MSP or not, had a very negative S score. As shown in Fig 11C, plates showed a regional reduction of colony sizes for genes on Chr. II, whereas the crosses involving genes located on other chromosomes (not boxed), showed no difference of colony sizes between the WT query plate and the chs1 query plate. In double mutants of chs1 combined with chs2-DAmP, fig1, fat1, cst26, or qdr3, we amplified by genomic PCR all genes and intergenic regions starting from CHS2 up to QDR3 and found that there were no rearrangements or deletions present apart from the intended single gene deletion. Interestingly, cst26 is one of the gene deletions sitting in the middle of the chromosomal region where deletions show negative S scores with chs1 (Fig 11A). As mentioned above, elo2 and elo3 also have very negative S scores in combination with cst26, similar to chs1 (Fig 11A). In this case however only very few genes immediately adjacent to cst26 show a negative S score with elo2 or elo3. The phenomenon of regionally concentrated negative interactions shown in Fig 11A is not an isolated phenomenon, since several such regions can easily be identified on a heat map of SPLOS Genetics | DOI:10.1371/journal.pgen.July 27,17 /Yeast E-MAP for Identification of Membrane Transporters Operating Lipid Flip FlopFig 11. Chs1-interacting cluster on chromosome II. (A) correlation and selected interaction heat maps of a series of linked genes on the right arm of Chr. II, deletion of which causes significant negative interactions with chs1. Genes are in the same order as they are situated along the chromosome. (B) colonies present on MSP-E-MAP plates after the last selection of double mutants combining deletions in genes indicated on top of column and indicated to the left. As indicated, double mutant colonies are from crosses with chs1 used as query or being on an array. Tda5 was added as a neutral control, having an S score of 0.07 with chs1. (C) Shows plates from quadruplicate crosses of either a WT query (upper image) or chs1 query strain (lower image) with an array plate containing many genes from different chromosomes including all the genes from YBR020W to YBR078W on the right arm of Chr. II. All genes in this chromosomal region are boxed, in red if there is a size difference visible to the naked eye between the WT and chs1 plate, in yellow if there isn't. Boxes are dotted for MSP genes that are part of the E-MAP set and are shown in panel A. doi:10.1371/journal.pgen.1006160.gscores where the genes are ordered according to their chromosomal location as shown in Fig 12. As the order in this matrix clusters each gene with the genes that sit next to it on the chromosome, all the irrelevant very negative interactions generated by proximity of two deletions on a same chromosome (< 100 kb) and hence marked with grey dots are clustering along the diagonal (Fig 12). Uniform interactions of all deletions in certain chromosomal regions withPLOS Genetics | DOI:10.1371/journal.pgen.July 27,18 /Yeast E-MAP for Identification of Membrane Transporters Operating Lipid Flip FlopFig 12. Interaction score heat map with genes ordered according to their chromosomal location. Standard names were replaced by the systematic names in the MSP-E-MAP interaction matrix (S3A Table) and.
