Human telomerase RNA and box H/ACA scaRNAs share a common Cajal body–specific localization signal

Telomerase is a ribonucleoprotein reverse transcriptase that uses its RNA component as a template for synthesis of telomeric DNA repeats at the ends of linear eukaryotic chromosomes. Here, fluorescence in situ hybridization demonstrates that in HeLa cancer cells, human telomerase RNA (hTR) accumulates in the nucleoplasmic Cajal bodies (CBs). Localization of transiently expressed hTR to CBs is supported by a short sequence motif (411-UGAG-414) that is located in the 3′-terminal box H/ACA RNA-like domain of hTR and that is structurally and functionally indistinguishable from the CB-specific localization signal of box H/ACA small CB-specific RNAs. In synchronized HeLa cells, hTR shows the most efficient accumulation in CBs during S phase, when telomeres are most likely synthesized. CBs may function in post-transcriptional maturation (e.g., cap hypermethylation of hTR), but they may also play a role in the assembly and/or function of telomerase holoenzyme

Controlling Cellular P-TEFb Activity by the HIV-1 Transcriptional Transactivator Tat

The human immunodeficiency virus 1 (HIV-1) transcriptional transactivator (Tat) is essential for synthesis of full-length transcripts from the integrated viral genome by RNA polymerase II (Pol II). Tat recruits the host positive transcription elongation factor b (P-TEFb) to the HIV-1 promoter through binding to the transactivator RNA (TAR) at the 5′-end of the nascent HIV transcript. P-TEFb is a general Pol II transcription factor; its cellular activity is controlled by the 7SK small nuclear RNA (snRNA) and the HEXIM1 protein, which sequester P-TEFb into transcriptionally inactive 7SK/HEXIM/P-TEFb snRNP. Besides targeting P-TEFb to HIV transcription, Tat also increases the nuclear level of active P-TEFb through promoting its dissociation from the 7SK/HEXIM/P-TEFb RNP by an unclear mechanism. In this study, by using in vitro and in vivo RNA-protein binding assays, we demonstrate that HIV-1 Tat binds with high specificity and efficiency to an evolutionarily highly conserved stem-bulge-stem motif of the 5′-hairpin of human 7SK snRNA. The newly discovered Tat-binding motif of 7SK is structurally and functionally indistinguishable from the extensively characterized Tat-binding site of HIV TAR and importantly, it is imbedded in the HEXIM-binding elements of 7SK snRNA. We show that Tat efficiently replaces HEXIM1 on the 7SK snRNA in vivo and therefore, it promotes the disassembly of the 7SK/HEXIM/P-TEFb negative transcriptional regulatory snRNP to augment the nuclear level of active P-TEFb. This is the first demonstration that HIV-1 specifically targets an important cellular regulatory RNA, most probably to promote viral transcription and replication. Demonstration that the human 7SK snRNA carries a TAR RNA-like Tat-binding element that is essential for the normal transcriptional regulatory function of 7SK questions the viability of HIV therapeutic approaches based on small drugs blocking the Tat-binding site of HIV TAR

Neural Cell Response to Nanostructured Biosensor Surfaces

AbstractIn our work we investigate the interaction of cells and nanotextured surfaces as a model of implanted device surface and living tissue interaction. We developed a maskless nanostructuring method, which can be integrated into our neural biosensor fabrication process. Morphology of the fabricated nanograss was characterised using SEM. The nanorods are 520-800nm in height and their density is 18-70/μm2. Electrochemical impedance spectroscopy and contact angles of different surfaces were measured. The specific surface area is 30 times larger than the reference. The contact-angle can be tuned. The samples will be tested in viability and adhesion assays using neural cell cultures

Dynamic association of human mRNP proteins with mitochondrial tRNAs in the cytosol

Cytoplasmic localization, stability, and translation of mRNAs are controlled by their dynamic association of numerous mRNA-binding (mRNP) proteins, including cold shock domain (CSD)-containing proteins, heterogeneous nuclear ribonucleoproteins (hnRNPs), and serine/arginine-rich (SR) proteins. Here, we demonstrate that the most abundant human mRNP protein, the CSD-containing Y-box-binding protein 1 (YBX1), the closely related YBX3 protein, and other mRNP proteins, such as SRSF1, SRSF2, SRSF3, hnRNP Al, and H, specifically and efficiently interact with overlapping sets of mitochondrial tRNAs (mt tRNAs). In vitro reconstitution and in vivo binding experiments show that YBX1 recognizes the D- and/or T-stem-loop regions of mt tRNAs through relying on the RNA-binding capacity of its CSD. Cell fractionation and in vivo RNA-protein cross-linking experiments demonstrate that YBX1 and YBX3 interact with mt tRNAs in the cytosol outside of mitochondria. Cell fractionation and fluorescence in situ hybridization experiments provide evidence that mitochondrial autophagy promotes the release of mt tRNAs from the mitochondria into the cytoplasm. Association of mRNP proteins with mt tRNAs is highly dynamic; it is rapidly increased upon transcription inhibition and decreased during apoptosis. Although the cytoplasmic function of mt tRNAs remains elusive, their dynamic interactions with key mRNA-binding proteins may influence cytoplasmic mRNA stability and/or translation

Human intron-encoded AluACA RNAs and telomerase RNA share a common element promoting RNA accumulation

Mammalian cells express hundreds of intron-encoded box H/ACA RNAs which fold into a common hairpin-hinge-hairpin-tail structure, interact with 4 evolutionarily conserved proteins, dyskerin, Nop10, Nhp2 and Gar1, and function mainly in RNA pseudouridylation. The human telomerase H/ACA RNA (hTR) directs telomeric DNA synthesis and it carries a 5-terminal domain encompassing the telomeric template sequence. The primary hTR transcript is synthesized from an independent gene by RNA polymerase II and undergoes 3 end processing controlled by the 3-terminal H/ACA domain. The apical stem-loop of the 3 hairpin of hTR carries a unique biogenesis-promoting element, the BIO motif that promotes hTR processing and RNP assembly. AluACA RNAs represent a distinct class of human H/ACA RNAs; they are processed from intronic Alu repetitive sequences. As compared to canonical H/ACA RNAs, the AluACA RNAs carry unusually short or long 5 hairpins and generally, they accumulate at low levels. Here, we demonstrate that the suboptimal 5 hairpins are responsible for the weak expression of AluACA RNAs. We also show that AluACA RNAs frequently carry a processing/stabilization element that is structurally and functionally indistinguishable from the hTR BIO motif. Both hTR and AluACA biogenesis-promoting elements are located in the terminal stem-loop of the 3-terminal H/ACA hairpin, they show perfect structural conservation and are functionally interchangeable in in vivo RNA processing reactions. Our results demonstrate that the BIO motif, instead of being confined to hTR, is a more general H/ACA RNP biogenesis-facilitating element that can also promote processing/assembly of intron-encoded AluACA RNPs

Differential signal sensitivities can contribute to the stability of multispecies bacterial communities.

BACKGROUND: Bacterial species present in multispecies microbial communities often react to the same chemical signal but at vastly different concentrations. The existence of different response thresholds with respect to the same signal molecule has been well documented in quorum sensing which is one of the best studied inter-cellular signalling mechanisms in bacteria. The biological significance of this phenomenon is still poorly understood, and cannot be easily studied in nature or in laboratory models. The aim of this study is to establish the role of differential signal response thresholds in stabilizing microbial communities. RESULTS: We tested binary competition scenarios using an agent-based model in which competing bacteria had different response levels with respect to signals, cooperation factors or both, respectively. While in previous scenarios fitter species outcompete slower growing competitors, we found that stable equilibria could form if the fitter species responded to a higher chemical concentration level than the slower growing competitor. We also found that species secreting antibiotic could form a stable community with other competing species if antibiotic production started at higher response thresholds. CONCLUSIONS: Microbial communities in nature rely on the stable coexistence of species that necessarily differ in their fitness. We found that differential response thresholds provide a simple and elegant way for keeping slower growing species within the community. High response thresholds can be considered as self-restraint of the fitter species that allows metabolically useful but slower growing species to remain within a community, and thereby the metabolic repertoire of the community will be maintained. REVIEWERS: This article was reviewed by Michael Gromiha, Sebastian Maurer-Stroh, István Simon and L. Aravind

The Hsp90 chaperone controls the biogenesis of L7Ae RNPs through conserved machinery

RNA-binding proteins of the L7Ae family are at the heart of many essential ribonucleoproteins (RNPs), including box C/D and H/ACA small nucleolar RNPs, U4 small nuclear RNP, telomerase, and messenger RNPs coding for selenoproteins. In this study, we show that Nufip and its yeast homologue Rsa1 are key components of the machinery that assembles these RNPs. We observed that Rsa1 and Nufip bind several L7Ae proteins and tether them to other core proteins in the immature particles. Surprisingly, Rsa1 and Nufip also link assembling RNPs with the AAA + adenosine triphosphatases hRvb1 and hRvb2 and with the Hsp90 chaperone through two conserved adaptors, Tah1/hSpagh and Pih1. Inhibition of Hsp90 in human cells prevents the accumulation of U3, U4, and telomerase RNAs and decreases the levels of newly synthesized hNop58, hNHP2, 15.5K, and SBP2. Thus, Hsp90 may control the folding of these proteins during the formation of new RNPs. This suggests that Hsp90 functions as a master regulator of cell proliferation by allowing simultaneous control of cell signaling and cell growth