Dynein is a microtubule motor protein, involved in many cellular processes, including intracellular transport and cell division. LIS-1, a microtubule-binding protein, binds two sites in dynein: the cargo binding region and the motor domain. Although the significance of this interaction is unknown, loss of LIS-1 function phenocopies loss of the cytoplasmatic dynein heavy chain (DHC-1) function in C. elegans, and mutations in either gene cause lethality. Finally,
lis-1 loss of function causes changes in the filamentous actin (F-actin) distribution in migrating neurons (Kholmanskikh S. et al., J. Neurosci 2003). Surprisingly, we discovered that a temperature sensitive (ts) mutation of
dhc-1(
or195ts) also causes defects in the actin cytoskeleton. Therefore, we have characterized the LIS-1-dynein interaction in C. elegans and studied its effects on the actin cytoskeleton. To investigate the interactions between
dhc-1 and
lis-1 we have focused on the C. elegans germline. Compared to the wild type (N2),
lis-1 and
dhc-1 mutants displayed severe defects in the germline actin cytoskeleton, indicating that both
dhc-1 and
lis-1 function in cytoskeleton organization. To examine the double mutant loss of function phenotype we used RNAi to knock-down
lis-1 in
dhc-1(
or195ts) animals. Surprisingly, all actin cytoskeletal defects were rescued and the double-mutant animals produced viable progeny. To better understand the rescue phenomenon, we investigated dynein localization in embryos at the one-cell stage. In
dhc-1(
or195ts) mutants, dynein localized exclusively to the minus end of microtubules (close to or at MTOC), while in embryos rescued by knock-down of
lis-1 a substantial pool of dynein was redistributed back along microtubules. These results indicate that
lis-1 plays a crucial role in dynein function and localization. To determine whether additional genes are able to rescue
dhc-1(lf)-induced lethality, we assembled a list of candidate dynein-interacting genes (Zhong and Sternberg, Science 311, 2006). We tested 243 candidate genes for RNAi rescue of the
dhc-1(
or195ts) embryonic lethal phenotype at 25C. We found that genes which significantly suppressed
dhc-1 lethality were microtubule related (e.g.
lis-1) or actin related (e.g.
cap-1,
cap-2), as well as prefoldin subunits (subunit 5, subunit 1, subunit 4). These findings suggest that the integrity of the cytoskeleton may mediate rescue of
dhc-1(
or195ts). The effect of knock-down of these genes in wild-type and
dhc-1(
or195ts) backgrounds will be presented at the meeting.