Leveraging HILIC/ERLIC Separations for Online Nanoscale LC-MS/MS Analysis of Phosphopeptide Isoforms from RNA Polymerase II C-terminal Domain.

The eukaryotic RNA polymerase II (Pol II) multi-protein complex transcribes mRNA and coordinates several steps of co-transcriptional mRNA processing and chromatin modification. The largest Pol II subunit, Rpb1, has a C-terminal domain (CTD) comprising dozens of repeated heptad sequences (Tyr1-Ser2-Pro3-Thr4-Ser5-Pro6-Ser7), each containing five phospho-accepting amino acids. The CTD heptads are dynamically phosphorylated, creating specific patterns correlated with steps of transcription initiation, elongation, and termination.

Cohesin dysfunction results in cell wall defects in budding yeast.

Cohesin is a conserved chromatin-binding multisubunit protein complex involved in diverse chromosomal transactions such as sister-chromatid cohesion, chromosome condensation, regulation of gene expression, DNA replication, and repair. While working with a budding yeast temperature-sensitive mutant, mcd1-1, defective in a cohesin subunit, we observed that it was resistant to zymolyase, indicating an altered cell wall organization. The budding yeast cell wall is a strong but elastic structure essential for maintenance of cell shape and protection from extreme environmental challenges.

A SIR-independent role for cohesin in subtelomeric silencing and organization.

Cohesin is a key determinant of chromosome architecture due to its DNA binding and tethering ability. Cohesin binds near centromeres and chromosome arms and also close to telomeres, but its role near telomeres remains elusive. In budding yeast, transcription within 20 kb of telomeres is repressed, in part by the histone-modifying silent information regulator (SIR) complex.

Interaction of the Saccharomyces cerevisiae RING-domain protein Nse1 with Nse3 and the Smc5/6 complex is required for chromosome replication and stability.

Genomic stability is maintained by the concerted actions of numerous protein complexes that participate in chromosomal duplication, repair, and segregation. The Smc5/6 complex is an essential multi-subunit complex crucial for repair of DNA double-strand breaks. Two of its subunits, Nse1 and Nse3, are homologous to the RING-MAGE complexes recently described in human cells.

Interplay between Top1 and Mms21/Nse2 mediated sumoylation in stable maintenance of long chromosomes.

Genetic information in cells is encrypted in DNA molecules forming chromosomes of varying sizes. Accurate replication and partitioning of chromosomes in the crowded cellular milieu is a complex process involving duplication, folding and movement. Longer chromosomes may be more susceptible to mis-segregation or DNA damage and there may exist specialized physiological mechanisms preventing this.