On NuMA and nuclear (re)organization

The eukaryotic cell nucleus stars in a major cellular process, the expression of the genome it harbors. Nuclear organization is dynamic, different among cell types and physiological states, and intimately linked to the regulation of gene expression. NuMA participates in the formation, stabilization, and orientation of the mitotic spindle. Additionally, it has been proposed that NuMA is involved in nuclear organization, both in the assembly of nuclei upon mitosis and during interphase. However, this idea has been controversial and never thoroughly investigated. In the present study we undertook a multidisciplinary approach to investigate the potential role of NuMA as an organizer of nuclear structure. Bioinformatics analysis of NuMA distal C-terminal sequence (NuMA-CTDP, residues 1915-2095) revealed orthologous sequences in vertebrates and sequence homology to chromatin-associated proteins. Moreover, NuMA-CTDP appears to have a primarily beta fold structure present in protein regions involved in ligand-receptor interactions. Expression of NuMA residues 1965-2101, treatment with anti-NuMA C-terminus antibodies, or siRNA silencing of NuMA induced reorganization of chromatin in differentiated and differentiating nonneoplastic human mammary epithelial HMT-3522 S1 cells (HMECs). Based on these results, the possibility of NuMA participating in ATP-dependent chromatin remodeling, one major mechanism for chromatin (re)organization, was investigated. NuMA cofractionated, coimmunoprecipitated, and colocalized with the ATPase SNF2H (present in several chromatin remodeling complexes) in HMECs. Mass spectrometry analysis from HeLa cells revealed other chromatin-associated proteins of interest copurifying with NuMA, some of which were further assessed in HMECs. This thesis demonstrates that NuMA may have a role in nuclear structure by organizing chromatin. Possible specific functions of NuMA in nuclear organization in relation to chromatin remodeling and the regulation of gene expression are discussed