The Role of Olfactory Cues in the Sequential Radiation of a Gall-boring Beetle, Mordellistena convicta

1. Herbivorous insects often have close associations with specific host plants, and their preferences for mating and ovipositing on a specific host-plant species can reproductively isolate populations, facilitating ecological speciation. Volatile emissions from host plants can play a major role in assisting herbivores to locate their natal host plants and thus facilitate assortative mating and host-specific oviposition. 2. The present study investigated the role of host-plant volatiles in host fidelity and oviposition preference of the gall-boring, inquiline beetle, Mordellistena convicta LeConte (Coleoptera: Mordellidae), using Y-tube olfactometers. Previous studies suggest that the gall-boring beetle is undergoing sequential host-associated divergence by utilising the resources that are created by the diverging populations of the gall fly, Eurosta solidaginis Fitch (Diptera: Tephritidae), which induces galls on the stems of goldenrods including Solidago altissima L. (Asteraceae) and Solidago gigantea Ait. 3. Our results show that M. convicta adults are attracted to galls on their natal host plant, avoid the alternate host galls, and do not respond to volatile emissions from their host-plant stems. 4. These findings suggest that the gall-boring beetles can orient to the volatile chemicals from host galls, and that beetles can use them to identify suitable sites for mating and/or oviposition. Host-associated mating and oviposition likely play a role in the sequential radiation of the gall-boring beetle.

Alba-domain proteins of Trypanosoma brucei are cytoplasmic RNA-binding proteins that interact with the translation machinery

Trypanosoma brucei and related pathogens transcribe most genes as polycistronic arrays that are subsequently processed into monocistronic mRNAs. Expression is frequently regulated post-transcriptionally by cis-acting elements in the untranslated regions (UTRs). GPEET and EP procyclins are the major surface proteins of procyclic (insect midgut) forms of T. brucei. Three regulatory elements common to the 3' UTRs of both mRNAs regulate mRNA turnover and translation. The glycerol-responsive element (GRE) is unique to the GPEET 3' UTR and regulates its expression independently from EP. A synthetic RNA encompassing the GRE showed robust sequence-specific interactions with cytoplasmic proteins in electromobility shift assays. This, combined with column chromatography, led to the identification of 3 Alba-domain proteins. RNAi against Alba3 caused a growth phenotype and reduced the levels of Alba1 and Alba2 proteins, indicative of interactions between family members. Tandem-affinity purification and co-immunoprecipitation verified these interactions and also identified Alba4 in sub-stoichiometric amounts. Alba proteins are cytoplasmic and are recruited to starvation granules together with poly(A) RNA. Concomitant depletion of all four Alba proteins by RNAi specifically reduced translation of a reporter transcript flanked by the GPEET 3' UTR. Pulldown of tagged Alba proteins confirmed interactions with poly(A) binding proteins, ribosomal protein P0 and, in the case of Alba3, the cap-binding protein eIF4E4. In addition, Alba2 and Alba3 partially cosediment with polyribosomes in sucrose gradients. Alba-domain proteins seem to have exhibited great functional plasticity in the course of evolution. First identified as DNA-binding proteins in Archaea, then in association with nuclear RNase MRP/P in yeast and mammalian cells, they were recently described as components of a translationally silent complex containing stage-regulated mRNAs in Plasmodium. Our results are also consistent with stage-specific regulation of translation in trypanosomes, but most likely in the context of initiation.

The role of olfactory cues in the sequential radiation of a gall-boring beetle

Species diversity itself may cause additional species diversity. According to recent findings, some species modify their environment in such a way that they facilitate the creation of new niches for other species to evolve to fill. Given the vast speciesdiversity of insects, the occurrence of such sequential radiation of species is likely common among herbivorous insects and the species that depend on them, many of them being insects as well. Herbivorous insects often have close associations with specific host plants and their preferences for mating and ovipositing on a specific host-plant species can reproductively isolate host-specific populations, facilitating speciation. Previous research by our laboratory has established that there are two distinct populations of thegall fly, Eurosta solidaginis (Tephritidae), which attack different species of goldenrods, Solidago altissima (Asteraceae) and S. gigantea. The gall fly’s host-associated differentiation is facilitating the divergence and potential speciation of twosubpopulations of the gall-boring beetle Mordellistena convicta (Mordellidae) by providing new resources (galls on stems of the galdenrods) for the gall-boring beetles. These beetles exist as two host-plant associated populations of inquilines that inhabit the galls induced by the gall fly. While our previous research has provided genetic and behavioral evidence for host-race formation, little is known about the role of their host plants in assortative mating and oviposition-site selection of the gall-boring beetles’ hostassociated populations. Volatile emissions from host plants can play a major role in assisting herbivores to locate their natal host plants and thus facilitate assortative mating and host-specific oviposition. The present study investigated the role of host-plant volatiles in host fidelity (mating on the host plant) and oviposition preference of M. convicta by measuring its behavioral responses to the host-plant volatile emissions using Y-tube olfactometers. In total, we tested behavioral responses of 615 beetles. Our resultsshow that M. convicta adults are attracted to their natal host galls (67% of S. altissima-emerging beetles and 70% of S. gigantea-emerging beetles) and avoid the alternate host galls (75% of S. altissima-emerging beetles and 66% of S. gigantea-emerging beetles),while showing no preference for, or avoidance of, ungalled plants from either species. This suggests that the gall beetles can orient to the volatile chemicals emitted by the galls and can potentially use them to identify suitable sites for mating and/or oviposition. Thus, host-associated mating and oviposition may play a role in the sequential speciation of the gall-boring beetle.

Integrity and regeneration of mechanotransduction machinery regulate aminoglycoside entry and sensory cell death

Sound perception requires functional hair cell mechanotransduction (MET) machinery, including the MET channels and tip-link proteins. Prior work showed that uptake of ototoxic aminoglycosides (AG) into hair cells requires functional MET channels. In this study, we examined whether tip-link proteins, including Cadherin 23 (Cdh23), regulate AG entry into hair cells. Using time-lapse microscopy on cochlear explants, we found rapid uptake of gentamicin-conjugated Texas Red (GTTR) into hair cells from three-day-old Cdh23(+/+) and Cdh23(v2J/+) mice, but failed to detect GTTR uptake in Cdh23(v2J/v2J) hair cells. Pre-treatment of wildtype cochleae with the calcium chelator 1,2-bis(o-aminophenoxy) ethane-N,N,N',N'-tetraacetic acid (BAPTA) to disrupt tip-links also effectively reduced GTTR uptake into hair cells. Both Cdh23(v2J/v2J) and BAPTA-treated hair cells were protected from degeneration caused by gentamicin. Six hours after BAPTA treatment, GTTR uptake remained reduced in comparison to controls; by 24 hours, drug uptake was comparable between untreated and BAPTA-treated hair cells, which again became susceptible to cell death induced by gentamicin. Together, these results provide genetic and pharmacologic evidence that tip-links are required for AG uptake and toxicity in hair cells. Because tip-links can spontaneously regenerate, their temporary breakage offers a limited time window when hair cells are protected from AG toxicity.

When small non-coding RNAs meet the ribosome: Tuning the translational machinery

The functions of ribosomes in translation are complex and involve different types of activities critical for decoding the genetic code, linkage of amino acids via amide bonds to form polypeptide chains, as well as the release and proper targeting of the synthesized protein. Non-protein-coding RNAs (ncRNAs) have been recognized to be crucial in establishing regulatory networks.1 However all of the recently discovered ncRNAs involved in translation regulation target the mRNA rather than the ribosome. The main goal of this project is to identify potential novel ncRNAs that directly bind and possibly regulate the ribosome during protein biosynthesis. To address this question we applied various stress conditions to the archaeal model organism Haloferax volcanii and deep-sequenced the ribosome-associated small ncRNA interactome. In total we identified 6.250 ncRNA candidates. Significantly, we observed the emersed presence of tRNA-derived fragments (tRFs). These tRFs have been identified in all domains of life and represent a growing, yet functionally poorly understood, class of ncRNAs. Here we present evidence that tRFs from H. volcanii directly bind to ribosomes. In the presented genomic screen of the ribosome-associated RNome a 26 residue long fragment originating from the 5’ part of valine tRNA was by far the most abundant tRF. The Val-tRF is processed in a stress- dependent manner and was found to primarily target the small ribosomal subunit in vitro and in vivo. As a consequence of ribosome binding, Val-tRF reduces protein synthesis by interfering with peptidyl transferase activity. Therefore this tRF functions as ribosome-bound small ncRNA capable of regulating gene expression in H. volcanii under environmental stress conditions probably by fine-tuning the rate of protein production.2 Currently we are investigating the binding site of this tRF on the 30S subunit in more detail.

An infectious bat-derived chimeric influenza virus harbouring the entry machinery of an influenza A virus.

In 2012, the complete genomic sequence of a new and potentially harmful influenza A-like virus from bats (H17N10) was identified. However, infectious influenza virus was neither isolated from infected bats nor reconstituted, impeding further characterization of this virus. Here we show the generation of an infectious chimeric virus containing six out of the eight bat virus genes, with the remaining two genes encoding the haemagglutinin and neuraminidase proteins of a prototypic influenza A virus. This engineered virus replicates well in a broad range of mammalian cell cultures, human primary airway epithelial cells and mice, but poorly in avian cells and chicken embryos without further adaptation. Importantly, the bat chimeric virus is unable to reassort with other influenza A viruses. Although our data do not exclude the possibility of zoonotic transmission of bat influenza viruses into the human population, they indicate that multiple barriers exist that makes this an unlikely event.