HOT1 is a mammalian direct telomere repeat‐binding protein contributing to telomerase recruitment

Telomeres are repetitive DNA structures that, together with the shelterin and the CST complex, protect the ends of chromosomes. Telomere shortening is mitigated in stem and cancer cells through the de novo addition of telomeric repeats by telomerase. Telomere elongation requires the delivery of the telomerase complex to telomeres through a not yet fully understood mechanism. Factors promoting telomerase–telomere interaction are expected to directly bind telomeres and physically interact with the telomerase complex. In search for such a factor we carried out a SILAC‐based DNA–protein interaction screen and identified HMBOX1, hereafter referred to as homeobox telomere‐binding protein 1 (HOT1). HOT1 directly and specifically binds double‐stranded telomere repeats, with the in vivo association correlating with binding to actively processed telomeres. Depletion and overexpression experiments classify HOT1 as a positive regulator of telomere length. Furthermore, immunoprecipitation and cell fractionation analyses show that HOT1 associates with the active telomerase complex and promotes chromatin association of telomerase. Collectively, these findings suggest that HOT1 supports telomerase‐dependent telomere elongation.     

Moderate calcium channel dysfunction in adult mice with inducible cardiomyocyte-specific excision of the cacnb2 gene

The major L-type voltage-gated calcium channels in heart consist of an α1C (Ca(V)1.2) subunit usually associated with an auxiliary β subunit (Ca(V)β2). In embryonic cardiomyocytes, both the complete and the cardiac myocyte-specific null mutant of Ca(V)β2 resulted in reduction of L-type calcium currents by up to 75%, compromising heart function and causing defective remodeling of intra- and extra-embryonic blood vessels followed by embryonic death. Here we conditionally excised the Ca(V)β2 gene (cacnb2) specifically in cardiac myocytes of adult mice (KO). Upon gene deletion, Ca(V)β2 protein expression declined by >96% in isolated cardiac myocytes and by >74% in protein fractions from heart. These latter protein fractions include Ca(V)β2 proteins expressed in cardiac fibroblasts. Surprisingly, mice did not show any obvious impairment, although cacnb2 excision was not compensated by expression of other Ca(V)β proteins or changes of Ca(V)1.2 protein levels. Calcium currents were still dihydropyridine-sensitive, but current density at 0 mV was reduced by <29%. The voltage for half-maximal activation was slightly shifted to more depolarized potentials in KO cardiomyocytes when compared with control cells, but the difference was not significant. In summary, Ca(V)β2 appears to be a much stronger modulator of L-type calcium currents in embryonic than in adult cardiomyocytes. Although essential for embryonic survival, Ca(V)β2 down-regulation in cardiomyocytes is well tolerated by the adult mice.     

Origins and population genetics of weedy red rice in the USA

Weedy red rice (Oryza sativa spontonea) is a persistent and problematic weed of rice culture worldwide. A major hypothesis for the mechanism of production of this weed in South and Southeast Asia is hybridization between cultivated rice (Oryza sativa) and wild rice (Oryza rufipogon). However, weedy red rice can often be found outside the range of O. rufipogon leaving questions on the origin and process behind weedy rice infestations. In the USA, weedy red rice was first documented as early as 1846 and has continued to affect rice production areas. In this study, we attempt to identify the origin and population structure of weedy red rice sampled from the USA using both DNA sequence data from a neutral nuclear locus as well as microsatellite genotype data. Results suggest that two major accessions of weedy rice exist, strawhull and blackhull, and these forms may both hybridize with the cultivated rice of the USA, O. sativa japonica. Using population assignment of multilocus genotype signatures with principal component analysis and structure, an Asian origin is supported for US weedy rice. Additionally, hybridization between strawhull and blackhull varieties was inferred and may present the opportunity for the production of new weedy forms in the future.     

The N-Terminal Domain of cGAS Determines Preferential Association with Centromeric DNA and Innate Immune Activation in the Nucleus

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A human-Tetrahymena pseudoknot chimeric telomerase RNA reconstitutes a nonprocessive enzyme in vitro that is defective in telomere elongation

The phylogenetically-derived secondary structures of telomerase RNAs (TR) from ciliates, yeasts and vertebrates are surprisingly conserved and contain a pseudoknot domain at a similar location downstream of the template. As the pseudoknot domains of Tetrahymena TR (tTR) and human TR (hTR) mediate certain similar functions, we hypothesized that they might be functionally interchangeable. We constructed a chimeric TR (htTR) by exchanging the hTR pseudoknot sequences for the tTR pseudoknot region. The chimeric RNA reconstituted human telomerase activity when coexpressed with hTERT in vitro, but exhibited defects in repeat addition processivity and levels of DNA synthesis compared to hTR. Activity was dependent on tTR sequences within the chimeric RNA. htTR interacted with hTERT in vitro and dimerized predominantly via a region of its hTR backbone, the J7b/8a loop. Introduction of htTR in telomerase-negative cells stably expressing hTERT did not reconstitute an active enzyme able to elongate telomeres. Thus, our results indicate that the chimeric RNA reconstituted a weakly active nonprocessive human telomerase enzyme in vitro that was defective in telomere elongation in vivo. This suggests that there may be species-specific requirements for pseudoknot functions.     

Distribution patterns and bioerosion of the sea urchin Centrostephanus coronatus (Diadematoida: Diadematidae), at the reef of Playa Blanca, Colombian Pacific

Regular sea-urchins are one of the main bioeroding organisms affecting coral reefs around the world. The abundance, distribution and bioerosion rate of the sea-urchin Centrostephanus coronatus, were determined in different reef zones of Playa Blanca fringing reef (Gorgona Island, Colombian pacific coast) during 1997 and 1998. The erosion rates were determined calcinating the gut content of the sea-urchins to eliminate all organic components and preserve the inorganic portion of calcium carbonate. C. coronatus showed the highest densities towards the central zones of the reef (plain-crest and front) (12.4 ind/m2; range 0-48 ind/m2). The highest mean bioerosion rate was 0.103 kgCaCO3/m2/yr in the reef plain-crest (0-0.69 kgCaCO3/m2/yr). In the other zones, (back reef and reef front) the mean bioerosion rates were 0.071 (range 0-0.39) and 0.052 (range 0-0.31) kgCaCO3/m2/yr respectively. According to the present data, it can be seen that the destruction of coralline skeletons, produced in this reef by sea-urchins is rather low, compared with the abrasion caused by these organisms in other places of the world. However, the combined action of C. coronatus and other bioeroding organisms (borers and grazers). along with some adverse environmental factors to corals, can be causing a negative balance between normal processes of reef accretion-destruction in Gorgona Island reefs.     

Distribution of endogenous albumin across the rat aortic wall as revealed by quantitative immunocytochemistry

Endogenous albumin was revealed over thin sections of rat aortic wall, with high resolution and specificity, by applying the protein A-gold immunocytochemical technique. Gold particles, revealing albumin antigenic sites, were observed over plasmalemmal vesicles in endothelial cells and over the interstitial space throughout the thickness of the aortic wall. The distribution of the labeling in the interstitial space varied from region to region and was associated with the collagen fibers, following the orientation of the bundles. The morphometric evaluation of this labeling demonstrated a first peak in labeling intensity in the intima followed by a steep decrease with low levels in the media, and an increasing gradient towards the adventitia. In the subendothelium, a moderate labeling was observed at the base of the endothelial cells of both aortic and capillary endothelia, followed by a decreasing gradient. Ratios between the labeling density in the intima as well as in the adventitia and that in the capillary lumen (plasma albumin) revealed different concentrations of albumin in these compartments. Endogenous albumin, under steady-state conditions, is thus unevenly distributed over the interstitial spaces across the rat aortic wall, and appears associated along the collagen fibers.