The nuclear pore complex is a macrostructure within the nuclear envelope (NE) composed of multiple copies of around 32 different proteins called nucleoporins. Nucleoporins interact with each other to form in some cases subcomplexes, such as the Nup107-160 subcomplex, consisting of 7-9 different nucleoporins. The Nup107-160 subcomplex plays important roles in nucleocytoplasmic transport, NE assembly and kinetochore function. However, the molecular mechanisms underlying these functions and the roles of the individual subcomplex members remain elusive. To dissect the function of the Nup107-160 subcomplex we are analysing two alleles of Nup107/npp-5 (
ok1966,
tm3039).
npp-5 mutants display temperature-dependent embryonic and larval lethality; 6% of homozygous
tm3039 F2 mutants develop into fertile adults at 20 degrees Celsius whereas all mutants die at 25 degrees Celsius. Surprisingly, NPP-5 is dispensable for NE recruitment of all other nucleoporins tested and nuclear protein import appears unaffected in the absence of NPP-5. In contrast, NPP-5 is essential for proper kinetochores localisation of NPP-15, another member of the Nup107-160 subcomplex, whereas recruitment of NPP-10C and MEL-28 to the kinetochores is NPP-5-independent. We found that
tm3039 mutants are hypersensitive to anoxia, suggesting that the spindle assembly checkpoint (SAC) could be compromised. Strikingly, down-regulation of SAC protein Mad1/MDF-1 by RNAi causes 100% embryonic lethality and severe DNA segregation defects in
tm3039 embryos, suggesting that the SAC needs to be activated for the survival of
tm3039 embryos. In support of this, kinetochore protein HIM-10 and Aurora kinase AIR-2 are less abundant on mitotic chromatin in
tm3039 embryos. However, we do not observe kinetochore recruitment of MDF-1 or Mad2/MDF-2 in
tm3039 embry os, but we found that interphase accumulation of MDF-1 at the NE is strictly dependent on NPP-5. We are currently testing if NPP-5 interacts physically with MDF-1.