Deciphering the role of LIN-53 in centromere function in caenorhabditis elegans

Abstract

Every cell must segregate its genetic materials equally to its daughter cells. Errors in chromosome segregation can lead to aneuploidy, as observed in chromosomal abnormality diseases such as Down syndrome. Increase rates of chromosome missegregation can result in chromosome instability, as commonly seen in solid tumors. The centromere is the single, specialized chromosomal domain responsible for directing chromosome segregation. All functional centromeres contain a histone H3 variant called the centromere protein A (CENP-A), which is thought to be the epigenetic mark of the centromere. However, the underlying mechanism for CENP-A chromatin assembly is not fully understood. In Caenorhabditis elegans holocentromeres, only MIS18BP1/KNL-2, a CENP-A/HCP-3 localization factor, is known to be inter-dependent with CENP-A/HCP-3 for centromere localization. To further uncover the CENP-A/HCP-3 assembly pathway, we identified nucleosomal CENP-A/HCP-3-interacting proteins by immunoprecipitation followed by mass spectrometry (IP-MS), including a chromatin remodeler RbAp48/LIN-53. RbAp48/LIN-53 was known to bind to retinoblastoma (Rb) protein and is involved in many histone modifying complexes. Live cell imaging showed that after RbAp48/LIN-53 depletion, CENP-A/HCP-3 on mitotic chromosomes was reduced significantly, comparable to in MIS18BP1/KNL-2 depletion. RbAp48/LIN-53 depletion also resulted in mitotic defects such as anaphase bridges and chromosome missegregation, but not chromosome condensation defects. Surprisingly, RbAp48/LIN-53 depletion did not mislocalize MIS18BP1/KNL-2, suggesting RbAp48/LIN-53 does not act upstream of MIS18BP1/KNL-2 in the CENP-A/HCP-3 assembly process. RbAp48/LIN-53 may act as a histone chaperone for CENP-A/HCP-3. Currently, we are analyzing whether one of the seven RbAp48/LIN-53-containing histone modifying complexes is involved in this function. This study will enhance the understanding of the centromere assembly pathway, and will also shed light on the development of artificial chromosomes. Artificial chromosomes with functional centromeres can be designed for accurate transmission, which can be used as vectors for gene therapy purposes.