The Cdk5 Homologue, Crp, Regulates Endocytosis and Secretion in Dictyostelium and Is Necessary for Optimum Growth and Differentiation

Abstract

Dictyostelium Crp is a member of the cyclin-dependent kinase (Cdk) family of proteins. It is most related in sequence to mammalian Cdk5, which unlike other members of the family, has functions that are unrelated to the cell cycle. In order to better understand the function of Crp in Dictyostelium, we overexpressed a dominant negative form, Crp-D144N, under the control of the actin 15 promoter. Cells overexpressing Crp-D144N exhibit a reduced growth rate in suspension culture and reduced rates of fluid-phase endocytosis and phagocytosis. There is no reduction in Cdc2 kinase activity in extracts from cells overexpressing Crp-D144N, suggesting that the growth defect is not due to inhibition of Cdc2. In addition to the growth defect, the act15::crp-D144N transformants aggregate at a slower rate than wild-type cells and form large aggregation streams. These eventually break up to form small aggregates and most of these do not produce mature fruiting bodies. The aggregation defect is fully reversed in the presence of wild-type cells but terminal differentiation is only partially rescued. In act15::crp-D144N transformants, the countin component of the counting factor, a secreted protein complex that regulates the breakup of streams, mostly appears outside the cell as degradation products and the reduced level of the intact protein may at least partially account for the initial formation of the large aggregation streams. Our observations indicate that Crp is important for both endocytosis and efflux and that defects in these functions lead to reduced growth and aberrant development.

Bestatin Inhibits Cell Growth, Cell Division, and Spore Cell Differentiation in Dictyostelium discoideum

“…PsaA is a binding partner of Cdk5 (17). D. discoideum cells expressing a dominant negative form of Cdk5 grow 3 times slower and produce 80% less spores than the wild type (46). Treatment of cells with roscovitine, an inhibitor of Cdks, also inhibits D. discoideum growth and spore cell differentiation (18).…”

“…Thus, NumA1 likely regulates the cell cycle and plays a role in pstO cell differentiation. Cdk5 is a nucleoplasmic and cytoplasmic protein that regulates growth and spore cell differentiation in D. discoideum (17,18,46).…”

The cyclin‐dependent kinase inhibitor roscovitine inhibits kinase activity, cell proliferation, multicellular development, and Cdk5 nuclear translocation in Dictyostelium discoideum

“…1C,D). Cdk5 activity had previously been shown to be required for optimal cell proliferation in Dictyostelium [Sharma et al, 2002]. Therefore, a strain over-expressing a Cdk5-GFP fusion protein [AX3/ [act15]:cdk5:GFP; Huber and O'Day, 2011a] was used to analyze the effect of Cdk5-GFP over-expression on roscovitine-inhibited axenic growth.…”

“…Several studies have linked Cdk5 dysregulation to advanced melanoma and neurodegenerative diseases such as amyotrophic lateral sclerosis and Alzheimer's disease [Dhavan and Tsai, 2001; Lee et al, 2010;Abdullah et al, 2011;Crews et al, 2011]. In Dictyostelium, Cdk5 has been shown to be required for optimal growth and differentiation, however, attempts to generate a knockout mutant have been unsuccessful [Sharma et al, 2002]. A recent study showed that Cdk5 localizes to both the nucleus and cytoplasm in Dictyostelium and binds to puromycin-sensitive aminopeptidase A [PsaA; Huber and O'Day, 2011a].…”

Orphan kinases turn eccentric

“…86 PFTAIREs emerged in metazoans and are present in species that lack a nervous system, e.g., in demosponges. In contrast, true PCTAIRE kinases ("CDK16"-homologs) are only present in animals, again supporting the hypothesis that these kinases might have an important role in neurons.…”