Mars promotes dTACC dephosphorylation on mitotic spindles to ensure spindle stability

Abstract

Microtubule-associated proteins (MAPs) ensure the fidelity of chromosome segregation by controlling microtubule (MT) dynamics and mitotic spindle stability. However, many aspects of MAP function and regulation are poorly understood in a developmental context. We show that mars, which encodes a Drosophila melanogaster member of the hepatoma up-regulated protein family of MAPs, is essential for MT stabilization during early embryogenesis. As well as associating with spindle MTs in vivo, Mars binds directly to protein phosphatase 1 (PP1) and coimmunoprecipitates from embryo extracts with minispindles and Drosophila transforming acidic coiled-coil (dTACC), two MAPs that function as spindle assembly factors. Disruption of binding to PP1 or loss of mars function results in elevated levels of phosphorylated dTACC on spindles. A nonphosphorylatable form of dTACC is capable of rescuing the lethality of mars mutants. We propose that Mars mediates spatially controlled dephosphorylation of dTACC, which is critical for spindle stabilization.

Mars, aDrosophilaprotein related to vertebrate HURP, is required for the attachment of centrosomes to the mitotic spindle during syncytial nuclear divisions

“…Consistent with this, we did not detect any localized staining for Mars in the mars 91 homozygous mutant embryos. A second recently published null allele of mars causes phenotypes essentially identical to the ones we report here, but these embryos never develop beyond the fifth nuclear division cycle (Tan et al, 2008). Whether this apparent discrepancy in the lethality of the two alleles is caused by some minor residual function still preserved in the mars 91 allele or by some differences in the genetic background of both alleles remains to be shown.…”

RanGTP is required for meiotic spindle organization and the initiation of embryonic development in Drosophila

“…Mars is the Drosophila homolog of HURP, a spindle assembly factor regulated by RanGTP (Wilde, 2006) and has been shown to have a role in the attachment of the centrosome to the mitotic spindle during Drosophila embryogenesis (Tan et al, 2008;Yang and Fan, 2008;Zhang et al, 2009). To study the role of Mars in meiotic spindle assembly, we examined mar 1 , which is a null allele that deletes part of the coding region (Tan et al, 2008). Like ran T24N mutants, mature oocytes homozygous for the mars 1 mutation were able to assemble a bipolar spindle but failed to properly taper the microtubules at the poles (Fig.…”

“…2E). To test whether these similarities could be the result of ran T24N mutant oocytes failing to activate Mars, we stained ran T24N mutant oocytes with an antibody against Mars (Tan et al, 2008). In wildtype or ran + -expressing oocytes, Mars colocalized with tubulin, except at the spindle poles and the central spindle (Fig.…”

“…Further studies are needed, however, because the role of Aurora A in Drosophila female meiosis is not known. In addition, Mars might have a role in promoting the dephosphorylation of TACC (Tan et al, 2008). Overall, RanGTP might have a specific role in organizing spindle poles but might not be required for chromosome-promoted spindle assembly in oocytes.…”

“…The primary antibodies were rat anti-SUB antibody (used at 1:75) (Jang et al, 2005), rat anti-HA (Roche, clone 3F10: 1:25), rat anti-INCENP (1:500) (Wu et al, 2008), mouse anti-RCC1 (1:20) (Frasch, 1991), rabbit anti-RanGAP (1:800) (Kusano et al, 2001), rabbit anti-Mars (Tan et al, 2008) and mouse anti-Lamin Dm 0 (1:800) (Klapper et al, 1997). These primary antibodies were detected with either a Cy3 or Cy5 secondary antibody preabsorbed against a range of mammalian serum proteins (Jackson Labs) and Drosophila embryos.…”