A novel probe for phosphatidylinositol 4-phosphate reveals multiple pools beyond the Golgi

Polyphosphoinositides are an important class of lipid that recruit specific effector proteins to organelle membranes. One member, phosphatidylinositol 4-phosphate (PtdIns4P) has been localized to Golgi membranes based on the distribution of lipid binding modules from PtdIns4P effector proteins. However, these probes may be biased by additional interactions with other Golgi-specific determinants. In this paper, we derive a new PtdIns4P biosensor using the PtdIns4P binding of SidM (P4M) domain of the secreted effector protein SidM from the bacterial pathogen Legionella pneumophila. PtdIns4P was necessary and sufficient for localization of P4M, which revealed pools of the lipid associated not only with the Golgi but also with the plasma membrane and Rab7-positive late endosomes/lysosomes. PtdIns4P distribution was determined by the localization and activities of both its anabolic and catabolic enzymes. Therefore, P4M reports a wider cellular distribution of PtdIns4P than previous probes and therefore will be valuable for dissecting the biological functions of PtdIns4P in its assorted membrane compartments.     

Genetic insights into the functional elements of language

Language disorders cover a wide range of conditions with heterologous and overlapping phenotypes and complex etiologies harboring both genetic and environmental influences. Genetic approaches including the identification of genes linked to speech and language phenotypes and the characterization of normal and aberrant functions of these genes have, in recent years, unraveled complex details of molecular and cognitive mechanisms and provided valuable insight into the biological foundations of language. Consistent with this approach, we have reviewed the functional aspects of allelic variants of genes which are currently known to be either causally associated with disorders of speech and language or impact upon the spectrum of normal language ability. We have also reviewed candidate genes associated with heritable speech and language disorders. In addition, we have evaluated language phenotypes and associated genetic components in developmental syndromes that, together with a spectrum of altered language abilities, manifest various phenotypes and offer details of multifactorial determinants of language function. Data from this review have revealed a predominance of regulatory networks involved in the control of differentiation and functioning of neurons, neuronal tracks and connections among brain structures associated with both cognitive and language faculties. Our findings, furthermore, have highlighted several multifactorial determinants in overlapping speech and language phenotypes. Collectively this analysis has revealed an interconnected developmental network and a close association of the language faculty with cognitive functions, a finding that has the potential to provide insight into linguistic hypotheses defining in particular, the contribution of genetic elements to and the modular nature of the language faculty.     

Transmissibility of the Monkeypox Virus Clades via Respiratory Transmission: Investigation Using the Prairie Dog-Monkeypox Virus Challenge System

Monkeypox virus (MPXV) is endemic within Africa where it sporadically is reported to cause outbreaks of human disease. In 2003, an outbreak of human MPXV occurred in the US after the importation of infected African rodents. Since the eradication of smallpox (caused by an orthopoxvirus (OPXV) related to MPXV) and cessation of routine smallpox vaccination (with the live OPXV vaccinia), there is an increasing population of people susceptible to OPXV diseases. Previous studies have shown that the prairie dog MPXV model is a functional animal model for the study of systemic human OPXV illness. Studies with this model have demonstrated that infected animals are able to transmit the virus to naive animals through multiple routes of exposure causing subsequent infection, but were not able to prove that infected animals could transmit the virus exclusively via the respiratory route. Herein we used the model system to evaluate the hypothesis that the Congo Basin clade of MPXV is more easily transmitted, via respiratory route, than the West African clade. Using a small number of test animals, we show that transmission of viruses from each of the MPXV clade was minimal via respiratory transmission. However, transmissibility of the Congo Basin clade was slightly greater than West African MXPV clade (16.7% and 0% respectively). Based on these findings, respiratory transmission appears to be less efficient than those of previous studies assessing contact as a mechanism of transmission within the prairie dog MPXV animal model.     

Evidence for targeting common siRNA hotspots and GC preference by plant Dicer-like proteins

Small interfering (si)RNAs isolated from Brassica juncea leaves infected by Turnip mosaic virus (TuMV) were characterized by cloning and sequencing. The TuMV siRNA population was dominated by 21 and 22-nt long species originated mainly from the same siRNA hotspots, indicating operational similarity between the plant Dicer-like (DCL) enzymes. Robust GC bias was observed for TuMV siRNAs versus the virus genome, indicating that DCL was more likely to target GC-rich regions. Furthermore, dicot micro-(mi)RNAs displayed higher GC% than their DCL1 substrate RNAs, implicating that the GC bias may be ancient, therefore may be important for the RNAi technology.     

Visualization and manipulation of plasma membrane-endoplasmic reticulum contact sites indicates the presence of additional molecular components within the STIM1-Orai1 Complex

STIM1, a recently identified endoplasmic reticulum (ER) protein, rapidly translocates to a plasma membrane-adjacent ER compartment upon depletion of the ER Ca(2+) stores. Here we use a novel means, namely a chemically inducible bridge formation between the plasma and ER membranes, to highlight the plasma membrane-adjacent ER compartment and show that this is the site where STIM1 and its Ca(2+) channel partner, Orai1, form a productive interaction upon store depletion. By changing the length of the linkers connecting the plasma and ER membranes, we show that Orai1 requires a larger space than STIM1 between the two membranes. This finding suggests that Orai1 is part of a larger macromolecular cluster with an estimated 11-14-nm protrusion to the cytoplasm, whereas the cytoplasmic domain of STIM1 fits in a space calculated to be less than 6 nm. We finally show that agonist-induced translocation of STIM1 is rapidly reversible and only partially affects STIM1 in the juxtanuclear ER compartment. These studies are the first to detect juxtaposed areas between the ER and the plasma membrane in live cells, revealing novel details of STIM1-Orai1 interactions.     

Effects of myeloid sirtuin 1 deficiency on hypothalamic neurogranin in mice fed a high-fat diet

Hypothalamic inflammation has been known as a contributor to high-fat diet (HFD)-induced insulin resistance and obesity. Myeloid-specific sirtuin 1 (SIRT1) deletion aggravates insulin resistance and hypothalamic inflammation in HFD-fed mice. Neurogranin, a calmodulin-binding protein, is expressed in the hypothalamus. However, the effects of myeloid SIRT1 deletion on hypothalamic neurogranin has not been fully clarified. To investigate the effect of myeloid SIRT1 deletion on food intake and hypothalamic neurogranin expression, mice were fed a HFD for 20 weeks. Myeloid SIRT1 knockout (KO) mice exhibited higher food intake, weight gain, and lower expression of anorexigenic proopiomelanocortin in the arcuate nucleus than WT mice. In particular, KO mice had lower ventromedial hypothalamus (VMH)-specific neurogranin expression. However, SIRT1 deletion reduced HFD-induced hypothalamic neurogranin. Furthermore, hypothalamic phosphorylated AMPK and parvalbumin protein levels were also lower in HFD-fed KO mice than in HFD-fed WT mice. Thus, these findings suggest that myeloid SIRT1 deletion affects food intake through VMH-specific neurogranin-mediated AMPK signaling and hypothalamic inflammation in mice fed a HFD.     

Molecular rearrangements of thylakoids after heavy metal poisoning, as seen by Fourier transform infrared (FTIR) and electron spin resonance (ESR) spectroscopy

The specific effects exerted by different heavy metals on both the function and the structure of the photosynthetic apparatus were addressed. The functional analysis performed via the fluorescence induction kinetics revealed that the applied toxic heavy metals can be classified into two groups: Cd and Ni had no significant effect on the photosynthetic electron transport, while Cu, Pb and Zn strongly inhibited the Photosystem II (PS II) activity, as evidenced by the dramatic decreases in both the variable (F_v) and the maximal (F_m) fluorescence. The structural effects of the heavy metal ions on the thylakoid membranes were considered in three relations: (1) lipids, (2) proteins — studied by Fourier transform infrared (FTIR) spectroscopy, and (3) lipid—protein interactions — investigated by electron spin resonance (ESR) spectroscopy using spin-labeled probe molecules. The studied heavy metal ions had only a non-specific rigidifying effect on the thylakoid lipids. As regards proteins, Cd and Ni had no effect on the course of their heat denaturation. The heat denaturation of the proteins was accompanied by a decrease in the -helix content (1656 cm^-1), a parallel increase in the disordered segments (1651 cm^-1), a decrease in the intramolecular -sheet (1636 cm^-1) content and the concomitant appearance of an intermolecular -structure (1621 cm^-1). In contrast with Cd and Ni, Cu and Zn blocked the appearance of the intermolecular -structure. Pb represented an intermediate case. It seems that these heavy metals alter the native membrane structure in such a way that heat-induced aggregation becomes more limited. The ESR data revealed that certain heavy metals also affect the lipid—protein interactions. While Cd and Ni had hardly any effect on the solvation fraction of thylakoid lipids, Cu, Pb and Zn increased the fraction of lipids solvating the proteins. On the basis of the FTIR and ESR data, it seems that Cu, Pb, and Zn increase the surfaces available for lipid—protein interactions by dissociating membrane protein complexes, and that these lipidated proteins have a smaller chance to aggregate upon heat denaturation. The data presented here indicate that the damaging effects of poisonous heavy metals are element-specific, Cu, Pb and Zn interact directly with the thylakoid membranes of the photosynthetic apparatus, while Cd and Ni interfere rather with other metabolic processes of plants.