Aedes aegypti mosquitoes are the primary vectors of numerous viruses that impact human health. As manipulation of reproduction has been proposed to suppress mosquito populations, elucidation of biological processes that enable males and females to successfully reproduce is necessary. One essential process is female sperm storage in specialized structures called spermathecae. Aedes aegypti females typically mate once, requiring them to maintain sperm viably to fertilize eggs they lay over their lifetime. Spermathecal gene products are required for Drosophila sperm storage and sperm viability, and a spermathecal-derived heme peroxidase is required for long-term Anopheles gambiae fertility. Products of the Ae. aegypti spermathecae, and their response to mating, are largely unknown. Further, although female blood-feeding is essential for anautogenous mosquito reproduction, the transcriptional response to blood-ingestion remains undefined in any reproductive tissue. We conducted an RNAseq analysis of spermathecae from unfed virgins, mated only, and mated and blood-fed females at 6, 24, and 72 h post-mating and identified significant differentially expressed genes in each group at each timepoint. A blood-meal following mating induced a greater transcriptional response in the spermathecae than mating alone. This study provides the first view of elicited mRNA changes in the spermathecae by a blood-meal in mated females.

Stu1 is the Saccharomyces cerevisiae member of the CLASP family of microtubule plus-end tracking proteins and is essential for spindle formation. A genomewide screen for gene deletions that are lethal in combination with the temperature-sensitive stu1-5 allele identified ldb18D. ldb18D cells exhibit defects in spindle orientation similar to those caused by a block in the dynein pathway. Consistent with this observation, ldb18D is synthetic lethal with mutations affecting the Kar9 spindle orientation pathway, but not with those affecting the dynein pathway. We show that Ldb18 is a component of dynactin, a complex required for dynein activity in yeast and mammalian cells. Ldb18 shares modest sequence and structural homology with the mammalian dynactin component p24. It interacts with dynactin proteins in two-hybrid and co-immunoprecipitation assays, and comigrates with them as a 20 S complex during sucrose gradient sedimentation. In ldb18D cells, the interaction between Nip100 (p150 Glued ) and Jnm1 (dynamitin) is disrupted, while the interaction between Jnm1 and Arp1 is not affected. These results indicate that p24 is required for attachment of the p150Glued arm to dynamitin and the remainder of the dynactin complex. The genetic interaction of ldb18D with stu1-5 also supports the notion that dynein/dynactin helps to generate a spindle pole separating force.

The organization of microtubules is determined in most cells by a microtubule-organizing center, which nucleates microtubule assembly and anchors their minus ends. In Saccharomyces cerevisiae cells lacking She1, cytoplasmic microtubules detach from the spindle pole body at high rates. Increased rates of detachment depend on dynein activity, supporting previous evidence that She1 inhibits dynein. Detachment rates are higher in G1 than in metaphase cells, and we show that this is primarily due to differences in the strengths of microtubule attachment to the spindle pole body during these stages of the cell cycle. The minus ends of detached microtubules are stabilized by the presence of γ-tubulin and Spc72, a protein that tethers the γ-tubulin complex to the spindle pole body. A Spc72–Kar1 fusion protein suppresses detachment in G1 cells, indicating that the interaction between these two proteins is critical to microtubule anchoring. Overexpression of She1 inhibits the loading of dynactin components, but not dynein, onto microtubule plus ends. In addition, She1 binds directly to microtubules in vitro, so it may compete with dynactin for access to microtubules. Overall, these results indicate that inhibition of dynein activity by She1 is important to prevent excessive detachment of cytoplasmic microtubules, particularly in G1 cells.

Background Mating induces behavioral and physiological changes in the arbovirus vector Aedes aegypti, including stimulation of egg development and oviposition, increased survival, and reluctance to re-mate with subsequent males. Transferred seminal fluid proteins and peptides derived from the male accessory glands induce these changes, though the mechanism by which they do this is not known. Results To determine transcriptome changes induced by seminal proteins, we injected extract from male accessory glands and seminal vesicles (MAG extract) into females and examined female lower reproductive tract (LRT) transcriptomes 24 h later, relative to non-injected controls. MAG extract induced 87 transcript-level changes, 31 of which were also seen in a previous study of the LRT 24 h after a natural mating, including 15 genes with transcript-level changes similarly observed in the spermathecae of mated females. The differentially-regulated genes are involved in diverse molecular processes, including immunity, proteolysis, neuronal function, transcription control, or contain predicted small-molecule binding and transport domains. Conclusions Our results reveal that seminal fluid proteins, specifically, can induce gene expression responses after mating and identify gene targets to further investigate for roles in post-mating responses and potential use in vector control.

Huntington's disease (HD) is a fatal progressive disease linked to expansion of glutamine repeats in the huntingtin protein and characterized by the progressive loss of cognitive and motor function. We show that expression of a mutant human huntingtin exon-1-GFP fusion construct results in nonspecific gene dysregulation that is significantly reduced by 50% due to coexpression of INT41, an intrabody specific for the proline-rich region of the huntingtin protein. Using stable PC12 cell lines expressing either inducible human mutant huntingtin (mHtt, Q73) or normal huntingtin (nHtt, Q23), we investigated the effect of rAAV6-INT41, an adeno-associated virus vector with the INT41 coding sequence, on the subcellular distribution of Htt. Compartmental fractionation 8 days after induction of Htt showed a 6-fold increased association of a dominate N-terminal mHtt fragment with DNA compared to N-terminal nHtt. Transduction with rAAV6-INT41 reduced DNA binding of N-terminal mHtt 6.5-fold in the nucleus and reduced nuclear translocation of the detected fragments. Subsequently, when rAAV6-INT41 is delivered to the striatum in the R6/2 mouse model, treated female mice exhibited executive function statistically indistinguishable from wild type, accompanied by reductions in Htt aggregates in the striatum, suggesting that rAAV6-INT41 is promising as a gene therapy for Huntington's disease.