Motor neuron function depends on neurotransmitter release from synaptic vesicles (SVs). Here we show that the UNC-4 homeoprotein and its transcriptional corepressor protein UNC-37 regulate SV protein levels in specific Caenorhabditis elegans motor neurons. UNC-4 is expressed in four classes (DA, VA, VC, and SAB) of cholinergic motor neurons. Antibody staining reveals that five different vesicular proteins (UNC-17, choline acetyltransferase, Synaptotagmin, Synaptobrevin, and RAB-3) are substantially reduced in unc-4 and unc-37 mutants in these cells; nonvesicular neuronal proteins (Syntaxin, UNC-18, and UNC-11) are not affected, however. Ultrastructural analysis of VA motor neurons in the mutant unc-4(e120) confirms that SV number in the presynaptic zone is reduced ( approximately 40%) whereas axonal diameter and synaptic morphology are not visibly altered. Because the UNC-4-UNC-37 complex has been shown to mediate transcriptional repression, we propose that these effects are performed via an intermediate gene. Our results are consistent with a model in which this unc-4 target gene ("gene-x") functions at a post-transcriptional level as a negative regulator of SV biogenesis or stability. Experiments with a temperature-sensitive unc-4 mutant show that the adult level of SV proteins strictly depends on unc-4 function during a critical period of motor neuron differentiation. unc-4 activity during this sensitive larval stage is also required for the creation of proper synaptic inputs to VA motor neurons. The temporal correlation of these events may mean that a common unc-4-dependent mechanism controls both the specificity of synaptic inputs as well as the strength of synaptic outputs for these motor neurons.

Mutations that inactivate the retinoblastoma (Rb) pathway are common in human tumors. Such mutations promote tumor growth by deregulating the G1 cell cycle checkpoint. However, uncontrolled cell cycle progression can also produce new liabilities for cell survival. To uncover such liabilities in Rb mutant cells, we performed a clonal screen in the Drosophila eye to identify second-site mutations that eliminate Rbf Ϫ cells, but allow Rbf ϩ cells to survive. Here we report the identification of a mutation in a novel highly conserved peptidyl prolyl isomerase (PPIase) that selectively eliminates Rbf Ϫ cells from the Drosophila eye.A N important goal of novel cancer therapy is to elicit mutations in the RB1 locus itself, but do carry mutations that target the pathway through the loss of cyclin-depenthe death of mutant tumor cells in the patient, while allowing normal cells to survive. The identification dent kinase (Cdk) inhibitors or overexpression of Cyclin D1 or Cdk4 (reviewed in Sherr and McCormick 2002). of gene products required for tumor cell survival can provide highly validated drug targets for the developAdditionally, the transforming activities of DNA tumor virus oncoproteins are mediated via their interaction ment of therapeutic inhibitors. Ideally, targets could be identified that would kill cancer cells while sparing with RB1 (Helt and Galloway 2003). The RB1 protein acts as a critical regulator of G1/S normal cells. A synthetic lethal screen is one method of identifying such targets. In this type of screen, cells phase progression by binding to members of the E2F are genetically altered to model tumor cells and one family of transcription factors (Dyson 1998; Nevins then screens for mutations that eliminate the model 2001). E2F-RB1 complexes prevent entry into S phase tumor cells but have little or no effect on wild-type cells.by actively repressing transcription through the recruitOne way to model tumor cells is to functionally inactiment of histone deacetylases and other chromatin modvate the RB1 gene. In addition to being mutated in ifiers to E2F-responsive promoters (Harbour and Dean retinoblastomas, where it was initially discovered, RB12000; Ogawa et al. 2002). Progression from G1 through is mutated in many other cancers including prostate S phase occurs when RB1 is inactivated through phos- ), bladder (Miyamoto et al. 1995, phorylation by the Cdk complexes Cyclin D/Cdk4 or parathyroid (Cryns et al. 1994), and 90% of small cell Cyclin D/Cdk6 and Cyclin E/Cdk2 (Lundberg and lung cancers (SCLCs) (Minna et al. 2002) Rbf is that it itself is required for embryonic survival.To circumvent this issue, we generated mosaic animals required for DNA synthesis and metabolism (Stevaux and Dyson 2002). In addition to its effects on cell prolifthat carry clones of Rbf Ϫ tissue in the eye, whereas the rest of the animal is Rbf ϩ . We then generated overlaperation, loss of RB1 predisposes cells to apoptosis through the actions of E2F on p53 (reviewed in Chau and Wang ping clones of homozygous induced muta...

No abstract