Tomato (Lycopersicon esculantum) ASR1 (abscisic acid stress ripening protein), a small plant-specific protein whose cellular mode of action defies deduction based on its sequence or homology analyses, is one of numerous plant gene products with unknown biological roles that become over-expressed under water- and salt-stress conditions. Steady-state cellular levels of tomato ASR1 mRNA and protein are transiently increased following exposure of plants to poly(ethylene glycol), NaCl or abscisic acid. Western blot and indirect immunofluorescence analysis with anti-ASR1 antibodies demonstrated that ASR1 is present both in the cytoplasmic and nuclear subcellular compartments; approx. one-third of the total ASR1 protein could be detected in the nucleus. Nuclear ASR1 is a chromatin-bound protein, and can be extracted with 1 M NaCl, but not with 0.5% Triton X-100. ASR1, overexpressed in Escherichia coli and purified to homogeneity, possesses zinc-dependent DNA-binding activity. Competitive-binding experiments and SELEX (systematic evolution of ligands by exponential enrichment) analysis suggest that ASR1 binds at a preferred DNA sequence.

Almost all animals contain mitochondria of maternal origin only, but the exact mechanisms underlying this phenomenon are still vague. We investigated the fate of Drosophila paternal mitochondria after fertilization. We demonstrate that the sperm mitochondrial derivative (MD) is rapidly eliminated in a stereotypical process dubbed paternal mitochondrial destruction (PMD). PMD is initiated by a network of vesicles resembling multivesicular bodies and displaying common features of the endocytic and autophagic pathways. These vesicles associate with the sperm tail and mediate the disintegration of its plasma membrane. Subsequently, the MD separates from the axoneme and breaks into smaller fragments, which are then sequestered by autophagosomes for degradation in lysosomes. We further provide evidence for the involvement of the ubiquitin pathway and the autophagy receptor p62 in this process. Finally, we show that the ubiquitin ligase Parkin is not involved in PMD, implying a divergence from the autophagic pathway of damaged mitochondria.

Patterning of the anterior-posterior body axis of the Drosophila embryo requires production of Nanos protein selectively in the posterior. Spatially restricted Nanos synthesis is accomplished by translational repression of unlocalized nanos mRNA together with translational activation of posteriorly localized nanos. Repression of unlocalized nanos mRNA is mediated by a bipartite translational control element (TCE) in its 3' untranslated region. TCE stem-loop II functions during embryogenesis, through its interaction with the Smaug repressor. Stem-loop III represses unlocalized nanos mRNA during oogenesis, but trans-acting factors that carry out this function have remained elusive. Here we identify a Drosophila hnRNP, Glorund, that interacts specifically with stem-loop III. We establish that the ability of the TCE to repress translation in vivo reflects its ability to bind Glorund in vitro. These data, together with the analysis of a glorund null mutant, reveal a specific role for an hnRNP in repression of nanos translation during oogenesis.

Caspases are executioners of apoptosis but also participate in a variety of vital cellular processes. Here, we identified Soti, an inhibitor of the Cullin-3-based E3 ubiquitin ligase complex required for caspase activation during Drosophila spermatid terminal differentiation (individualization). We further provide evidence that the giant inhibitor of apoptosis-like protein dBruce is a target for the Cullin-3-based complex, and that Soti competes with dBruce for binding to Klhl10, the E3 substrate recruitment subunit. We then demonstrate that Soti is expressed in a subcellular gradient within spermatids and in turn promotes proper formation of a similar dBruce gradient. Consequently, caspase activation occurs in an inverse graded fashion, such that the regions of the developing spermatid that are the last to individualize experience the lowest levels of activated caspases. These findings elucidate how the spatial regulation of caspase activation can permit caspase-dependent differentiation while preventing full-blown apoptosis.

ASR1 is a plant-specific, highly charged, low molecular weight polypeptide. Purified ASR1 was shown to posses sequence specific Zn 2+ + + + -dependent DNA binding activity (Kalifa et al . Biochemical Journal 381, 373-378, 2004). Steady-state levels of tomato Asr1 mRNA and protein are transiently increased following exposure of plants to polyethylene glycol, NaCl or abscisic acid. The biological role of ASR1 could not be deduced from sequence analyses or sequence homologies. Tobacco plants over-expressing tomato ASR1 have a decreased rate of water loss and improved salt tolerance. Upon exposure to salt, ASR1-over-expressing plants accumulate less Na + + + + and proline than wild-type plants, and also results in increased steadystate levels of other gene products under non-stressed plant growth conditions. Therefore, ASR1 is probably involved in the regulation of water-or salt-stress-modulated gene expression.