bims-micpro 2020-07-19 papers

Sci Adv. 2020 May;6(21): eaaz2059

A micropeptide encoded by lncRNA MIR155HG suppresses autoimmune inflammation via modulating antigen presentation.

Liman Niu, Fangzhou Lou, Yang Sun, Libo Sun, Xiaojie Cai, Zhaoyuan Liu, Hong Zhou, Hong Wang, Zhikai Wang, Jing Bai, Qianqian Yin, Junxun Zhang, Linjiao Chen, Danhong Peng, Zhenyao Xu, Yuanyuan Gao, Sibei Tang, Li Fan, Honglin Wang.

Many annotated long noncoding RNAs (lncRNAs) harbor predicted short open reading frames (sORFs), but the coding capacities of these sORFs and the functions of the resulting micropeptides remain elusive. Here, we report that human lncRNA MIR155HG encodes a 17-amino acid micropeptide, which we termed miPEP155 (P155). MIR155HG is highly expressed by inflamed antigen-presenting cells, leading to the discovery that P155 interacts with the adenosine 5'-triphosphate binding domain of heat shock cognate protein 70 (HSC70), a chaperone required for antigen trafficking and presentation in dendritic cells (DCs). P155 modulates major histocompatibility complex class II-mediated antigen presentation and T cell priming by disrupting the HSC70-HSP90 machinery. Exogenously injected P155 improves two classical mouse models of DC-driven auto inflammation. Collectively, we demonstrate the endogenous existence of a micropeptide encoded by a transcript annotated as "non-protein coding" and characterize a micropeptide as a regulator of antigen presentation and a suppressor of inflammatory diseases.

DOI: https://doi.org/10.1126/sciadv.aaz2059

Proteomics. 2020 Jul 15. e2000084

Complementarity of Different SDS-PAGE Gel Staining Methods for the Identification of Short Open Reading Frame-encoded Peptides.

Philipp T Kaulich, Liam Cassidy, Katrin Weidenbach, Ruth A Schmitz, Andreas Tholey.

Short open reading frame-encoded peptides (SEP) have been identified across all domains of life and are predicted to be involved in many biochemical processes, however, for the vast majority of SEP their biological function is still unknown. Optimized methodologies have to be used for the mass spectrometric analysis of SEP, because traditional methods of bottom-up proteomics show a bias against small proteins. Here, we investigated different staining methods for SDS-PAGE gels prior in-gel digestion following LC-MS/MS analysis for the identification of SEP in the archaeon Methanosarcina mazei. In total, we identified 45 SEP with at least one high confidence (FDR<1 %) unique peptide and five consecutive b- or y-ions in the MS2 spectrum. The staining methods provided complementary data. The highest number of SEP were identified in the samples stained with Coomassie brilliant blue. However, the highest quality of the identified SEP was achieved in the samples without staining. These comprehensive data sets demonstrate that in-gel digestion is well suited for the identification of SEP. This article is protected by copyright. All rights reserved.

Keywords: GeLC-MS; alternative ORF; microproteins; peptidomics; sORF

DOI: https://doi.org/10.1002/pmic.202000084

Front Microbiol. 2020 ;11 1384

Establishing Polycistronic Expression in the Model Microorganism Ustilago maydis.

Kira Müntjes, Magnus Philipp, Lisa Hüsemann, Nicole Heucken, Stefanie Weidtkamp-Peters, Kerstin Schipper, Matias D Zurbriggen, Michael Feldbrügge.

Eukaryotic microorganisms use monocistronic mRNAs to encode proteins. For synthetic biological approaches like metabolic engineering, precise co-expression of several proteins in space and time is advantageous. A straightforward approach is the application of viral 2A peptides to design synthetic polycistronic mRNAs in eukaryotes. During translation of these peptides the ribosome stalls, the peptide chain is released and the ribosome resumes translation. Thus, two independent polypeptide chains can be encoded from a single mRNA when a 2A peptide sequence is placed inbetween the two open reading frames. Here, we establish such a system in the well-studied model microorganism Ustilago maydis. Using two fluorescence reporter proteins, we compared the activity of five viral 2A peptides. Their activity was evaluated in vivo using fluorescence microscopy and validated using fluorescence resonance energy transfer (FRET). Activity ranged from 20 to 100% and the best performing 2A peptide was P2A from porcine teschovirus-1. As proof of principle, we followed regulated gene expression efficiently over time and synthesised a tri-cistronic mRNA encoding biosynthetic enzymes to produce mannosylerythritol lipids (MELs). In essence, we evaluated 2A peptides in vivo and demonstrated the applicability of 2A peptide technology for U. maydis in basic and applied science.

Keywords: 2A peptide; FRET; RNA transport; RRM; Ustilago maydis; mannosylerythritol lipid

DOI: https://doi.org/10.3389/fmicb.2020.01384