Understanding of the role of barrier-to-autointegration factor (BAF) in nuclear assembly, chromatin condensation and transcriptional regulation.

摘要

The nuclear envelope, composed of the inner and outer nuclear membranes, serves to compartmentalize the nucleus within the cell. LAP2, emerin and MAN1 are part of a family of proteins of the inner nuclear membrane that share a 43-residue LEM domain. The conserved LEM domain interacts with Barrier-to-autointegration factor (BAF), a small DNA-binding protein conserved in metazoans. Although BAF was first discovered as a cytosolic activity that prevented retroviral self-integration, its cellular function was unknown. To understand the role of BAF in the nucleus, I looked at the effect of BAF on nuclear assembly and chromatin decondensation in Xenopus nuclear extracts. This work showed that BAF has a potent effect on chromatin structure and nuclear assembly. I biochemically mapped the regions of interaction in BAF for emerin and DNA using a collection of 25 BAF mutants. The region of DNA interaction I determined coincided with structural predictions, while the region of emerin interaction in BAF complements the shape and hydrophobicity of the LEM domain. I also determined the null phenotype for BAF in C. elegans by RNA interference. Embryos depleted of BAF die by the 100-cell stage and have DNA segregation defects. Emerin and MAN1 are mislocalized in nuclei lacking BAF, while lamins appear disorganized. These findings suggest that BAF plays a role in the assembly and organization of LEM proteins and lamins. We also discovered a novel protein that is homologous to BAF, which we termed BAF-like (BAF-L). BAF-L is expressed in all tissues tested except for heart, skeletal muscle and kidney. BAF and BAF-L have distinct biochemical properties: BAF-L does not interact with emerin or DNA. However, BAF-L interacts with two regions in LAP2 and MAN1, with BAF, and with itself. A two-hybrid study with BAF-L as bait yielded a transcriptional regulator MLL3, as a potential partner. This discovery links BAF-L, and potentially BAF, to transcriptional regulatory complexes.