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The Structural Maintenance of Chromosome (SMC) complex, termed cohesin, is essential for sister chromatid cohesion. Cohesin is also important for chromosome condensation, DNA repair, and gene expression. Cohesin is comprised of Scc3, Mcd1, Smc1, and Smc3. Scc3 also binds Pds5 and Wpl1, cohesin-associated proteins that regulate cohesin function, and to the Scc2/4 cohesin loader. We mutagenized SCC3 to elucidate its role in cohesin function. A 5 amino acid insertion after Scc3 residue I358, or a missense mutation of residue D373 in the adjacent stromalin conservative domain (SCD) induce inviability and defects in both cohesion and cohesin binding to chromosomes. The I358 and D373 mutants abrogate Scc3 binding to Mcd1. These results define an Scc3 region extending from I358 through the SCD required for binding Mcd1, cohesin localization to chromosomes and cohesion. Scc3 binding to the cohesin loader, Pds5 and Wpl1 are unaffected in I358 mutant and the loader still binds the cohesin core trimer (Mcd1, Smc1 and Smc3). Thus, Scc3 plays a critical role in cohesin binding to chromosomes and cohesion at a step distinct from loader binding to the cohesin trimer. We show that residues Y371 and K372 within the SCD are critical for viability and chromosome condensation but dispensable for cohesion. However, scc3 Y371A and scc3 K372A bind normally to Mcd1. These alleles also provide evidence that Scc3 has distinct mechanisms of cohesin loading to different loci. The cohesion-competence, condensation-incompetence of Y371 and K372 mutants suggests that cohesin has at least one activity required specifically for condensation. [ABSTRACT FROM AUTHOR]

The Kaposi's sarcoma associated herpesvirus (KSHV) encoded LANA protein functions in latently infected cells as an essential participant in KSHV genome replication and as a driver of dysregulated cell growth. In a previous study, we have identified LANA interacting proteins using a protein array screen. Here, we explore the effect of LANA on the stability and activity of RLIM (RING-finger LIM-domain-interacting protein, encoded by the RNF12 gene) a novel LANA interacting protein identified in that protein screen. RLIM is an E3 ubiquitin ligase that leads to the ubiquitination and degradation of several transcription regulators, such as LMO2, LMO4, LHX2, LHX3, LDB1 and the telomeric protein TRF1. Expression of LANA leads to down-regulation of RLIM protein levels. This LANA-mediated RLIM degradation is blocked in the presence of the proteasome inhibitor, MG132. Therefore, the interaction between LANA and RLIM could be detected in co-immunoprecipitation assay only in the presence of MG132 to prevent RLIM degradation. A RING finger mutant RLIM (HH 590, 593 EE) is resistant to LANA mediated degradation, suggesting that LANA promotes RLIM auto-ubiquitination. Interestingly, we found that LANA enhanced the degradation of some RLIM substrates, such as LDB1 and LMO2, and prevented RLIM mediated degradation of others such as LHX3 and TRF1. We also show that transcription regulation by RLIM substrates is modulated by LANA. RLIM substrates are assembled into multi-protein transcription regulator complexes that regulate the expression of many cellular genes. Therefore, our study identified another way KSHV can modulate cellular gene expression. [ABSTRACT FROM AUTHOR]