The salience of the hues: colour cognition from an indigenous Australian perspective

Does natural language determine the way we think? If the so-called Sapir-Whorf hypothesis of linguistics were true then colour categorization would be an entirely arbitrary process dependant entirely on the language that we speak. For a while in academic circles this was the received wisdom: The fact that English had 11 colour terms and Dugum Dani (from Papua New Guinea) had only two was a factor attributed to the language in use. In 1969, Brent Berlin and Paul Kay ushered in the paradigm shift of Basic Colour Terms, a theory that defined the structure and evolution of colour terms in cultures (Berlin and Kay, 1969). According to this theory there was a neurophysiological basis for colour categorization, which implied that all human beings had the potential to see the same colours but naming the categories was an evolutionary process tied into the technological sophistication of a society. Language in other words played little or no part in colour perception. The initial theory was highly controversial with the demand growing for verification of the initial findings. To address the contentious issues surrounding Berlin and Kay’s theory, a project entitled the World Colour Survey was initiated with the goal of determining colour categorization patterns within 110 pre-literate cultures across the globe. This project has spanned more than 30 years with a definitive publication of the results still in the works. The current PhD project, which has been in progress since the dawn of time, involves an independent analysis and interpretation of the indigenous Australian component of the World Colour Survey raw data on colour categorization. It is both an exercise in secondary analysis as a research method and a tangential meditation on the ambiguity of knowledge that can be derived from extant data sets

An unbiased proteomics approach to identify human cytomegalovirus RNA-associated proteins

AbstractPost-transcriptional events regulate herpesvirus gene expression, yet few herpesvirus RNA-binding proteins have been identified. We used an unbiased approach coupling oligo(dT) affinity capture with proteomics to identify viral RNA-associated proteins during infection. Using this approach, we identified and confirmed changes in the abundance or activity of two host RNA-associated proteins, DHX9 and DDX3, in cells infected with human cytomegalovirus (HCMV). We also identified and confirmed previously unreported activities for the HCMV US22 and pp71 proteins as RNA-associated viral proteins and confirmed that a known viral RNA-binding protein, pTRS1, associates with RNA in infected cells. Further, we found that HCMV pp71 co-sedimented with polysomes, associated with host and viral RNAs, and stimulated the overall rate of protein synthesis. These results demonstrate that oligo(dT) affinity capture coupled with proteomics provides a rapid and straightforward means to identify RNA-associated viral proteins during infection that may participate in the post-transcriptional control of gene expression

The eIF4AIII RNA helicase is a critical determinant of human cytomegalovirus replication

Human cytomegalovirus (HCMV) was recently shown to encode a large number of spliced mRNAs. While the nuclear export of unspliced viral transcripts has been extensively studied, the role of host mRNA export factors in HCMV mRNA trafficking remains poorly defined. We found that the eIF4AIII RNA helicase, a component of the exon junction complex, was necessary for efficient virus replication. Depletion of eIF4AIII limited viral DNA accumulation, export of viral mRNAs from the nucleus, and the production of progeny virus. However eIF4AIII was dispensable for the association of viral transcripts with ribosomes. We found that pateamine A, a natural compound that inhibits both eIF4AI/II and eIF4AIII, has potent antiviral activity and inhibits HCMV replication throughout the virus lytic cycle. Our results demonstrate that eIF4AIII is required for efficient HCMV replication, and suggest that eIF4A family helicases may be a new class of targets for the development of host-directed antiviral therapeutics

An unbiased proteomics approach to identify human cytomegalovirus RNA-associated proteins

Post-transcriptional events regulate herpesvirus gene expression, yet few herpesvirus RNA-binding proteins have been identified. We used an unbiased approach coupling oligo(dT) affinity capture with proteomics to identify viral RNA-associated proteins during infection. Using this approach, we identified and confirmed changes in the abundance or activity of two host RNA-associated proteins, DHX9 and DDX3, in cells infected with human cytomegalovirus (HCMV). We also identified and confirmed previously unreported activities for the HCMV US22 and pp71 proteins as RNA-associated viral proteins and confirmed that a known viral RNA-binding protein, pTRS1, associates with RNA in infected cells. Further, we found that HCMV pp71 co-sedimented with polysomes, associated with host and viral RNAs, and stimulated the overall rate of protein synthesis. These results demonstrate that oligo(dT) affinity capture coupled with proteomics provides a rapid and straightforward means to identify RNA-associated viral proteins during infection that may participate in the post-transcriptional control of gene expression

Molecular basis for functional diversity among microbial Nep1-like proteins

Necrosis and ethylene-inducing peptide 1 (Nep1)-like proteins (NLPs) are secreted by several phytopathogenic microorganisms. They trigger necrosis in various eudicot plants upon binding to plant sphingolipid glycosylinositol phosphorylceramides (GIPC). Interestingly, HaNLP3 from the obligate biotroph oomycete Hyaloperonospora arabidopsidis does not induce necrosis. We determined the crystal structure of HaNLP3 and showed that it adopts the NLP fold. However, the conformations of the loops surrounding the GIPC headgroup-binding cavity differ from those of cytotoxic Pythium aphanidermatum NLPPya. Essential dynamics extracted from \u3bcs-long molecular dynamics (MD) simulations reveals a limited conformational plasticity of the GIPC-binding cavity in HaNLP3 relative to toxic NLPs. This likely precludes HaNLP3 binding to GIPCs, which is the underlying reason for the lack of toxicity. This study reveals that mutations at key protein regions cause a switch between nontoxic and toxic phenotypes within the same protein scaffold. Altogether, these data provide evidence that protein flexibility is a distinguishing trait of toxic NLPs and highlight structural determinants for a potential functional diversification of non-toxic NLPs utilized by biotrophic plant pathogens

Developmental bias in cleavage-stage mouse blastomeres

BACKGROUND: The cleavage-stage mouse embryo is composed of superficially equivalent blastomeres that will generate both the embryonic inner cell mass (ICM) and the supportive trophectoderm (TE). However, it remains unsettled whether the contribution of each blastomere to these two lineages can be accounted for by chance. Addressing the question of blastomere cell fate may be of practical importance, because preimplantation genetic diagnosis requires removal of blastomeres from the early human embryo. To determine whether blastomere allocation to the two earliest lineages is random, we developed and utilized a recombination-mediated, noninvasive combinatorial fluorescent labeling method for embryonic lineage tracing. RESULTS: When we induced recombination at cleavage stages, we observed a statistically significant bias in the contribution of the resulting labeled clones to the trophectoderm or the inner cell mass in a subset of embryos. Surprisingly, we did not find a correlation between localization of clones in the embryonic and abembryonic hemispheres of the late blastocyst and their allocation to the TE and ICM, suggesting that TE-ICM bias arises separately from embryonic-abembryonic bias. Rainbow lineage tracing also allowed us to demonstrate that the bias observed in the blastocyst persists into postimplantation stages and therefore has relevance for subsequent development. CONCLUSIONS: The Rainbow transgenic mice that we describe here have allowed us to detect lineage-dependent bias in early development. They should also enable assessment of the developmental equivalence of mammalian progenitor cells in a variety of tissues

Complex polar machinery required for proper chromosome segregation in vegetative and sporulating cells of Bacillus subtilis

Chromosome segregation is an essential process of cell multiplication. In prokaryotes, segregation starts with the newly replicated sister origins of replication, oriCs, which move apart to defined positions in the cell. We have developed a genetic screen to identify mutants defective in placement of oriC during spore development in the Gram-positive bacterium Bacillus subtilis. In addition to the previously identified proteins Soj and DivIVA, our screen identified several new factors involved in polar recruitment of oriC: a reported regulator of competence ComN, and the regulators of division site selection MinD and MinJ. Previous work implicated Soj as an important regulator of oriC positioning in the cell. Our results suggest a model in which the DivIVA-interacting proteins ComN and MinJ recruit MinD to the cell pole, and that these proteins work upstream of Soj to enable oriC placement. We show that these proteins form a polar complex, which acts in parallel with but distinct from the sporulation-specific RacA pathway of oriC placement, and also functions during vegetative growth. Our study further shows that MinD has two distinct cell cycle roles, in cell division and chromosome segregation, and highlights that cell probably use multiple parallel mechanisms to ensure accurate chromosome segregation.</p

Human cytomegalovirus pTRS1 stimulates cap-independent translation

Human cytomegalovirus (HCMV) manipulates multiple cellular processes to facilitate virus replication, including the control of mRNA translation. We previously showed that the HCMV TRS1 protein (pTRS1) promotes cap-dependent mRNA translation independent of its ability to antagonize the antiviral protein PKR. Here we find that pTRS1 enhances internal ribosome entry site (IRES) activity using a novel circular RNA reporter that lacks an mRNA cap and poly(A) tail. Additionally, pTRS1 expression increases the activity of cellular IRESs that control the expression of proteins needed for efficient HCMV replication. We find that the ability of pTRS1 to enhance cap-independent translation is separable from its ability to antagonize PKR, but requires the pTRS1 RNA binding domain. Together these data show that pTRS1 stimulates cap-independent translation and suggest a role for pTRS1 in alternative translation initiation pathways during HCMV infection

Differential Role for Host Translation Factors in Host and Viral Protein Synthesis during Human Cytomegalovirus Infection

The host eIF4F translation initiation complex plays a critical role the translation of capped mRNAs. Although human cytomegalovirus (HCMV) infection increases the abundance and activity of the host eIF4F complex, the requirement for eIF4F components in HCMV replication and mRNA translation has not been directly tested. In this study, we found that decreasing the abundance or activity of eIF4F from the start of infection inhibits HCMV replication. However, as infection progresses, viral mRNA translation and replication becomes increasingly resistant to eIF4F inhibition. During the late stage of infection the association of representative immediate-early, early, and late mRNAs with polysomes was not affected by eIF4F disruption. In contrast, eIF4F inhibition decreased the translation of representative host eIF4F-dependent mRNAs during the late stage of infection. A global analysis of the translation efficiency of HCMV mRNAs during the late stage of infection found that eIF4F disruption had a minimal impact on the association of HCMV mRNAs with polysomes but significantly diminished the translation efficiency of eIF4F-dependent host transcripts. Together, our data show that the translation of host eIF4F-dependent mRNAs remains dependent on eIF4F activity during HCMV infection. However, during the late stage of infection the translation efficiency of viral mRNAs does not correlate with the abundance or activity of the host eIF4F complex

Kinome Profiling Identifies Druggable Targets for Novel Human Cytomegalovirus (HCMV) Antivirals

Human cytomegalovirus (HCMV) is a significant cause of disease in immune-compromised adults and immune naïve newborns. No vaccine exists to prevent HCMV infection, and current antiviral therapies have toxic side effects that limit the duration and intensity of their use. There is thus an urgent need for new strategies to treat HCMV infection. Repurposing existing drugs as antivirals is an attractive approach to limit the time and cost of new antiviral drug development. Virus-induced changes in infected cells are often driven by changes in cellular kinase activity, which led us to hypothesize that defining the complement of kinases (the kinome), whose abundance or expression is altered during infection would identify existing kinase inhibitors that could be repurposed as new antivirals. To this end, we applied a kinase capture technique, multiplexed kinase inhibitor bead-mass spectrometry (MIB-MS) kinome, to quantitatively measure perturbations in >240 cellular kinases simultaneously in cells infected with a laboratory-adapted (AD169) or clinical (TB40E) HCMV strain. MIB-MS profiling identified time-dependent increases and decreases in MIB binding of multiple kinases including cell cycle kinases, receptor tyrosine kinases, and mitotic kinases. Based on the kinome data, we tested the antiviral effects of kinase inhibitors and other compounds, several of which are in clinical use or development. Using a novel flow cytometry-based assay and a fluorescent reporter virus we identified three compounds that inhibited HCMV replication with IC 50 values of 3 log decrease in virus replication. These results show the utility of MIB-MS kinome profiling for identifying existing kinase inhibitors that can potentially be repurposed as novel antiviral drugs