Orbit/Mast, the CLASP orthologue ofDrosophila, is required for asymmetric stem cell and cystocyte divisions and development of the polarised microtubule network that interconnects oocyte and nurse cells during oogenesis

Abstract

Drosophila oocyte differentiation is preceded by the formation of a polarised 16-cell cyst from a single progenitor stem cell as a result of four rounds of asymmetric mitosis followed by incomplete cytokinesis. We show that the Orbit/Mast microtubule-associated protein is required at several stages in the formation of such polarised 16-cell cysts. In wild-type cysts, the Orbit/Mast protein not only associates with the mitotic spindle and its poles, but also with the central spindle (spindle remnant), ring canal and fusome, suggesting it participates in interactions between these structures. In orbit mutants, the stem cells and their associated fusomes are eventually lost as Orbit/Mast protein is depleted. The mitotic spindles of those cystocytes that do divide are either diminutive or monopolar, and do not make contact with the fusome. Moreover, the spindle remnants and ring canals fail to differentiate correctly in such cells and the structure of fusome is compromised. The Orbit/Mast protein thus appears to facilitate multiple interactions of the fusome with mitotic spindles and ring canals. This ensures correct growth of the fusome into a branched asymmetrically distributed organelle that is pre-determinative of 16-cell cyst formation and oocyte fate specification. Finally the Orbit/Mast protein is required during mid-oogenesis for the organisation of the polarised microtubule network inside the 16-cell cyst that ensures oocyte differentiation. The localisation of CLIP-190 to such microtubules and to the fusome is dependent upon Orbit/Mast to which it is complexed.

Dynein and Mast/Orbit/CLASP have antagonistic roles in regulating kinetochore-microtubule plus-end dynamics

“…The same kind of dynein dependence was described for mammalian CLIP-170 (Dujardin et al, 1998;Tanenbaum et al, 2006). It was observed that in addition to dynein, CLIP-190 also binds Mast (Mathe et al, 2003). Since CLASPs are CLIP-associated-proteins that bind to CLIP-170 through its conserved C-terminal domain (the same domain that targets Mast to kinetochores) (Akhmanova et al, 2001), it was proposed that CLIP-170 might be a linker for dynein cargo (Dujardin et al, 1998).…”

Actin Cytoskeletal Organization in Drosophila Germline Ring Canals Depends on Kelch Function in a Cullin-RING E3 Ligase

“…We note that the ring canal phenotype seen in kelch mutants is remarkably specific: the loss-of-function ovarian germline phenotypes of thousands of genes-perhaps a third of the genome-have been examined (see Hudson and Cooley 2014 for review of systematic genetic screens performed in the ovary), and yet only four genes are known to result in a kelchlike phenotype. Two of these are kelch and Cul3, while in the other two, Impa2 and orbit, the phenotypes appear to result from a failure to localize Kelch to ring canals (Gorjanacz et al 2002;Mathe et al 2003). Thus to our knowledge, all characterized mutations that result in a kelch-like phenotype are required for Kelch function in some manner.…”

S. pombe CLASP needs dynein, not EB1 or CLIP170, to induce microtubule instability and slows polymerization rates at cell tips in a dynein-dependent manner

“…CLASPs are required to maintain a stem cell component in Drosophila and for ordered cell migration in vertebrates (Mathe et al 2003;Lee et al 2004) and can be critically important for localized modulation of microtubule dynamics (Inoue et al 2004). Mammalian CLASPs stabilize the association of microtubules and the cell cortex (Mimori-Kiyosue et al 2005).…”