## CELL SPECIFICITY IN YEAST MATING SWITCH

### CELL SPECIFICITY IN YEAST MATING SWITCH

8.2.         Cell specificity in yeast:

Mating occurs only between different types of cells but not between cells of the same mating type. The difference between ‘a and α’ is made due to the presence of different sets of genes which are actively expressed and suppressed in these two mating types. The different expression pattern of both mating types is due to MAT locus which is located on chromosome III in yeast. ‘MAT a’ and ‘MAT α’ are two different alleles of MAT locus.

In haploid cells, there is only one allele of MAT is present (‘MAT a’ or ‘MAT α’) that determine their mating type. The signals which are received by cell is responsible to start a signalling cascade. The ‘α cell’ have ‘MAT α’ locus, so in these cell two different proteins α1 and α2 expressed. α1 with MCM1 works as an activator of α specific proteins where α2 with MCM1 works to repress a specific gene. Haploid specific genes are also expressed and some of them work as the repressor of meiosis. So meiosis is blocked in haploid cells.

In 'a' type of cell ‘MAT a’ is present. But both α1 and α2 proteins are absent. So in a cell, MATa1 gene is expressed. Haploid specific genes are also expressed and some of them work as the repressor of meiosis. So meiosis is blocked in haploid cells. In a/α diploids possess both of the alleles of MAT locus (heterozygous for MAT locus). In these diploid cells, expression of MATα2 represses the ‘a specific’ genes and MATα2 /MATa1 dimer is formed which repress the haploid-specific genes. Meiosis is not blocked in diploid cells. Diploid cells undergo meiosis and form haploid spores again. The ‘a and α’ specific genes have loci for encoding mating factors, receptors and proteins for cell fusion.

8.3.         Regulation of cell specificity  in yeast:

The cell specification is a transcription control mechanism. Haploid cells express haploid-specific genes of their type. In a/α diploid cell both ‘a specific’ and ‘α specific’ genes are silent and diploid-specific genes are expressed. General transcription factor MCM1 with cell-specific transcription factors α1, α2 and a1 encoded by MAT locus regulate the cell type-specific transcription.

MCM is a member of the MADS family protein. (MADS proteins also involved in floral organs). MCM1 show different activities in haploid ‘a and α’ cells, it binds with α1 or α2 proteins.

8.4.         Activation of a specific gene:

In the yeast chromosome III, yeast mating type promoter have specific sequence named P-BOX which possesses MCM binding site. MCM1 binds to the P-box dimeric format a specific upstream regulatory sequence (URS) and stimulate the transcription of a specific gene.

Activation of ‘α specific’ genes:

The ‘α specific’ genes have 2 adjacent DNA sequences P-BOX and Q-BOX in their promoter region. MCM1 binds to the P-BOX and α1 with MCM1 bind to the Q-BOX in α specific URS and starts transcription from PQ site.

Gene repression: In a specific URS two α2 binding sites are also present. Binding of MCM and α2 simultaneously repress the transcription of a specific gene. MCM directs the DNA binding domains of α2 dimer toward α2 binding sites on a specific URS. So MCM1 play an important role to increase the affinity of the complex and thus play role in transcription regulation. If the number of α2 binding sites are present in the genome, it can regulate various protein expression. In diploid cell α2 form heterodimer with a1 which can repress both α1 and haploid-specific genes. The ‘specific’ genes are not expressed in ‘α specific’ and diploid cells.

The signalling pathway that induces mating:

Cells follow GPCR-FUS pathway. As mating factors bind to their receptors some proteins are expressed that cause cell cycle arrest in G1 and promote attachment and fusion of two haploid cells. Mating pheromones bind to their respective receptors and work, as the ligand. Binding of the signals (‘a or α’) to the receptor results in the activation of G protein by phosphorylation. Activated G protein dissociates from active Gβ-Gγ subunits which phosphorylate the various ‘Ste’ proteins like a cascade.

Activation of Ste4/Ste5  $\rightarrow$  Ste20  $\rightarrow$  Ste11 $\rightarrow$  Ste7

Ste 20, Ste 11, Ste 7 have MAP kinase activity. They phosphorylate the FUS3 protein.  Phosphorylated   FUS 3 start the transcription of MAT locus.

FUS3 also activate ‘Ste 12’ protein which works as a transcription factor. Fus-3 with SK-12 binds to pheromone responsive element (PRE) and transcribe the proteins, involved in mating, cell arrest, cell pairing, and fusion.

Ste12 bind on PRE when MCM1 bind on P-BOX. Binding of Ste12 on PRE stimulate the function of MCM1 as an activator.

The signalling mechanism is the same in both mating types of cells but the difference is in the receptor. The 'a' cells bear Ste receptor whereas ‘α cells’ have Ste3 receptor on their cell surface.

Mating switch:

In a colony of yeast where only one mating type is present (a type or α type), some cells switch their mating type and convert into opposite mating type (a to α or vice versa). This phenomenon is called a mating switch yeast can find the opposite mating type by mating switch process.

So if there are haploid cells of a single mating type found in a yeast colony. Mating type switching will cause cells of both ‘a and α’ mating type to be present in that population after some time.

8.5.         Cassette model for mating type switch:

This conversion of mating type become possible by a specific feature of yeast. The cell has two additional silent copies of MAT locus in a chromosome. That is HML (Hidden MAT Left) which is the silent copy of MAT α and HMR (Hidden MAT Right) silent copy of MAT a locus. These copies are not transcribed. SIR proteins bind with both HML and HMR form a heterochromatin scaffold and prevent the transcription of these regions. The HML and HMR are present near the heterochromatic region so cannot express itself due to Position effect variegation. Only the active MAT locus is transcribed.

The cassette model of mating type switch explains that switching is the result of the specific type of genetic recombination event which is initiated by cleavage of DNA sequence by HO endonuclease. HO gene is a haploid-specific gene (homothallic). Expression of this gene results in the production of HO endonuclease which cut the DNA at the MAT locus due to its specificity. As endonuclease cut the DNA, the exonuclease recognizes the ends and cut all the MAT sequence. The gap generated filled by the copying genetic information present at the HML or HMR. In this repair, a cell uses HML as a source of information.HML is just like MAT a locus. Thus HML locus is copied at MAT locus. This converts the mating type ‘a to α’ type. ‘α cells’ follow the same to convert in a type. So the expression of HO is crucial for the mating switch.

In yeast chromosome III Recombination Enhancers (RE) present on the left arm of the chromosome. In 'a' strain cell MCM1 binds to the enhancer, cause recombination of HML. Cell repairs the gap using HML and switches mating type to α. Whereas in case of α type α2 binds with RE thus RE suppressed and cell repair the gap by using HMR and convert ‘α’ type to ‘a’ type.