Evolutionary Divergence of Enzymatic Mechanisms for Tubulin Detyrosination.

The two related members of the vasohibin family, VASH1 and VASH2, encode human tubulin detyrosinases. Here we demonstrate that, in contrast to VASH1, which requires binding of small vasohibin binding protein (SVBP), VASH2 has autonomous tubulin detyrosinating activity. Moreover, we demonstrate that SVBP acts as a bona fide activator of both enzymes. Phylogenetic analysis of the vasohibin family revealed that regulatory diversification of VASH-mediated tubulin detyrosination coincided with early vertebrate evolution. Thus, as a model organism for functional analysis, we used Trypanosoma brucei (Tb), an evolutionarily early-branched eukaryote that possesses a single VASH and encodes a terminal tyrosine on both α- and ß-tubulin tails, both subject to removal. Remarkably, although detyrosination levels are high in the flagellum, TbVASH knockout parasites did not present any noticeable flagellar abnormalities. In contrast, we observed reduced proliferation associated with profound morphological and mitotic defects, underscoring the importance of tubulin detyrosination in cell division.

CSAP Acts as a Regulator of TTLL-Mediated Microtubule Glutamylation.

Tubulin glutamylation is a reversible posttranslational modification that accumulates on stable microtubules (MTs). While abnormally high levels of this modification lead to a number of disorders such as male sterility, retinal degeneration, and neurodegeneration, very little is known about the molecular mechanisms underlying the regulation of glutamylase activity. Here, we found that CSAP forms a complex with TTLL5, and we demonstrate that the two proteins regulate their reciprocal abundance. Moreover, we show that CSAP increases TTLL5-mediated glutamylation and identify the TTLL5-interacting domain. Deletion of this domain leads to complete loss of CSAP activating function without impacting its MT binding. Binding of CSAP to TTLL5 promotes relocalization of TTLL5 toward MTs. Finally, we show that CSAP binds and activates all of the remaining autonomously active TTLL glutamylases. As such, we present CSAP as a major regulator of tubulin glutamylation and associated functions.

Expression map of the human exome in CD34+ cells and blood cells: increased alternative splicing in cell motility and immune response genes.

BACKGROUND: Hematopoietic cells are endowed with very specific biological functions, including cell motility and immune response. These specific functions are dramatically altered during hematopoietic cell differentiation, whereby undifferentiated hematopoietic stem and progenitor cells (HSPC) residing in bone marrow differentiate into platelets, red blood cells and immune cells that exit into the blood stream and eventually move into lymphoid organs or inflamed tissues. The contribution of alternative splicing (AS) to these functions has long been minimized due to incomplete knowledge on AS events in hematopoietic cells. PRINCIPAL FINDINGS: Using Human Exon ST 1.0 microarrays, the entire exome expression profile of immature CD34+ HSPC and mature whole blood cells was mapped, compared to a collection of solid tissues and made freely available as an online exome expression atlas (Amazonia Exon! : http://amazonia.transcriptome.eu/exon.php). At a whole transcript level, HSPC strongly expressed EREG and the pluripotency marker DPPA4. Using a differential splicing index scheme (dsi), a list of 849 transcripts differentially expressed between hematopoietic cells and solid tissues was computed, that included NEDD9 and CD74. Some of these genes also underwent alternative splicing events during hematopoietic differentiation, such as INPP4B, PTPLA or COMMD6, with varied contribution of CD3+ T cells, CD19+ B cells, CD14+ or CD15+ myelomonocytic populations. Strikingly, these genes were significantly enriched for genes involved in cell motility, cell adhesion, response to wounding and immune processes. CONCLUSION: The relevance and the precision provided by this exon expression map highlights the contribution of alternative splicing to key feature of blood cells differentiation and function.