Glatiramer acetate treatment normalizes deregulated microRNA expression in relapsing remitting multiple sclerosis

The expression of selected microRNAs (miRNAs) known to be involved in the regulation of immune responses was analyzed in 74 patients with relapsing remitting multiple sclerosis (RRMS) and 32 healthy controls. Four miRNAs (miR-326, miR-155, miR-146a, miR-142-3p) were aberrantly expressed in peripheral blood mononuclear cells from RRMS patients compared to controls. Although expression of these selected miRNAs did not differ between treatment-naïve (n = 36) and interferon-beta treated RRMS patients (n = 18), expression of miR-146a and miR-142-3p was significantly lower in glatiramer acetate (GA) treated RRMS patients (n = 20) suggesting that GA, at least in part, restores the expression of deregulated miRNAs in MS.     

The CONJUDOR pipeline for multiplexed knockdown of gene pairs identifies RBBP-5 as a germ cell reprogramming barrier in C. elegans

Multiple gene activities control complex biological processes such as cell fate specification during development and cellular reprogramming. Investigating the manifold gene functions in biological systems requires also simultaneous depletion of two or more gene activities. RNA interference-mediated knockdown (RNAi) is commonly used in Caenorhabditis elegans to assess essential genes, which otherwise lead to lethality or developmental arrest upon full knockout. RNAi application is straightforward by feeding worms with RNAi plasmid-containing bacteria. However, the general approach of mixing bacterial RNAi clones to deplete two genes simultaneously often yields poor results. To address this issue, we developed a bacterial conjugation-mediated double RNAi technique ‘CONJUDOR’. It allows combining RNAi bacteria for robust double RNAi with high-throughput. To demonstrate the power of CONJUDOR for large scale double RNAi screens we conjugated RNAi against the histone chaperone gene lin-53 with more than 700 other chromatin factor genes. Thereby, we identified the Set1/MLL methyltransferase complex member RBBP-5 as a novel germ cell reprogramming barrier. Our findings demonstrate that CONJUDOR increases efficiency and versatility of RNAi screens to examine interconnected biological processes in C. elegans with high-throughput.     

Regulation of Argonaute Slicer Activity by Guide RNA 3′ End Interactions with the N-terminal Lobe

Structural studies indicate that binding of both the guide RNA (siRNA and miRNA) and the target mRNA trigger substantial conformational changes in the Argonaute proteins. Here we explore the role of the N-terminal lobe (and its PAZ domain) in these conformational changes using biochemical and cell culture-based approaches. In vitro, whereas deletion (or mutation) of the N-terminal lobe of DmAgo1 and DmAgo2 had no effect on binding affinity to guide RNAs, we observed a loss of protection of the 3′ end of the guide RNA and decreased target RNA binding; consistent with this, in cells, loss of function DmAgo1 PAZ variant proteins (PAZ6 and ΔN-PAZ) still bind RNA, although the RNAs are shorter than normal. We also find that deletion of the N-terminal lobe results in constitutive activation of endogenous PIWI domain-based cleavage activity in vitro, providing insights into how cleavage activity may be regulated in vivo in response to different types of pairing interactions with the target mRNAs.     

Nickel and iron incorporation into soluble hydrogenase of Alcaligenes eutrophus

Archives of Microbiology - Qualitative and quantitative determination of proteins of the soluble hydrogenase (hydrogen: NAD+ oxidoreductase, EC 1.12.1.2) from Alcaligenes eutrophus H16 was done by...