Growth Retardation, Polyploidy, and Multinucleation Induced by Clast3, a Novel Cell Cycle-regulated Protein

Abstract

We have identified a novel gene, Clast3, by subtraction of cDNAs derived from activated and naive B lymphocytes. Clast3 expression is elevated in cycling cells and down-regulated in cells undergoing growth arrest, indicating that its expression is controlled in a cell cycledependent manner. The deduced amino acid sequence of Clast3 cDNA exhibits no significant homology to the known proteins in mammalian and other species. Immunofluorescence staining revealed that Clast3 localizes into discrete nuclear foci. Forced expression of Clast3 results in growth retardation, polyploidy, and generation of multinucleated cells. Treatment of Clast3 transfectants with nocodazole, a spindle-damaging agent, greatly enhances the incidence of the multinucleated cells, suggesting that Clast3 overexpression impairs the same checkpoint activated by nocodazole. Down-regulation of Clast3 expression by antisense oligonucleotides results in a decrease of cells at G 2 -M phase and a concomitant increase of apoptotic cells. These findings indicate that Clast3 is a novel cell cycle-regulated protein and that its constitutive overexpression induces polyploidy and multinucleation by interfering with the mitotic spindle checkpoint.Genetic stability is achieved by the coordinated regulation of DNA replication and repair, chromosomal segregation, and cell cycle checkpoints (1, 2). There are several checkpoints that act to ensure the orderly progression of critical events in the cell cycle. The mitotic spindle checkpoint functions to delay the metaphase to anaphase transition until all pairs of sister chromatids are attached to spindle microtubes, thereby ensuring the correct segregation of duplicated chromosomes into the two daughter cells (3,4). A number of proteins have been identified that sense the kinetochore microtube attachment and regulate the separation of sister chromatids. These include the kinetochore-associated CENPE, MAD, and BUB proteins, the anaphase-promoting complex/cyclosome and its associated cofac-

Some assembly required: dedicated chaperones in eukaryotic proteasome biogenesis

“…An indication that this might be the case came from the discovery of a heterodimeric protein complex in mammals, called PAC1–PAC2 (for p roteasome a ssembling c haperone 1 and 2; Hirano et al, 2005). These proteins had been identified previously, but no connection to proteasome function had been made (Bahar et al, 2002; Possik et al, 2004; Vidal-Taboada et al, 2000). PAC1 and PAC2 were identified as proteins coprecipitating with an epitope-tagged proteasome subunit (Hirano et al, 2005).…”

Global Analysis of the Cortical Neuron Proteome

“…A number of hemopoietic proteins were identified such as mDomino, a myeloid cell differentiation factor (14), myeloid zinc finger protein-2, a zinc-finger transcription factor (15), and Clast3, a novel cell cycle-regulated protein first identified in activated and naive B lymphocytes (16). Unique transcriptional activators and repressors were identified in cortical neurons, such as myocardin-related transcription factor A and enhancer of rudimentary homologue.…”

ADD66, a Gene Involved in the Endoplasmic Reticulum-associated Degradation of α-1-Antitrypsin-Z in Yeast, Facilitates Proteasome Activity and Assembly

“…18, October 2007 3781 CLAST3 and HCCA3, a gene that is up-regulated in hepatic cancers (Wang et al, 2001;Bahar et al, 2002). Based on these observations and our data presented above, we examined whether 20S proteasome assembly intermediates accumulate in yeast lacking Add66p.…”

“…CLAST3 and HCCA3, a gene that is up-regulated in hepatic cancers (Wang et al, 2001;Bahar et al, 2002).…”