Functional analysis of RRAS2 pathogenic variants with a Noonan-like phenotype

Introduction: RRAS2, a member of the R-Ras subfamily of Ras-like low-molecular-weight GTPases, is considered to regulate cell proliferation and differentiation via the RAS/MAPK signaling pathway. Seven RRAS2 pathogenic variants have been reported in patients with Noonan syndrome; however, few functional analyses have been conducted. Herein, we report two patients who presented with a Noonan-like phenotype with recurrent and novel RRAS2 pathogenic variants (p.Gly23Val and p.Gly24Glu, respectively) and the results of their functional analysis.Materials and methods: Wild-type (WT) and mutant RRAS2 genes were transiently expressed in Human Embryonic Kidney293 cells. Expression of RRAS2 and phosphorylation of ERK1/2 were confirmed by Western blotting, and the RAS signaling pathway activity was measured using a reporter assay system with the serum response element-luciferase construct. WT and p.Gly23Val RRAS2 were expressed in Drosophila eye using the glass multiple reporter-Gal4 driver. Mutant mRNA microinjection into zebrafish embryos was performed, and the embryo jaws were observed.Results: No obvious differences in the expression of proteins WT, p.Gly23Val, and p.Gly24Glu were observed. The luciferase reporter assay showed that the activity of p.Gly23Val was 2.45 ± 0.95-fold higher than WT, and p.Gly24Glu was 3.06 ± 1.35-fold higher than WT. For transgenic flies, the p.Gly23Val expression resulted in no adults flies emerging, indicating lethality. For mutant mRNA-injected zebrafish embryos, an oval shape and delayed jaw development were observed compared with WT mRNA-injected embryos. These indicated hyperactivity of the RAS signaling pathway.Discussion: Recurrent and novel RRAS2 variants that we reported showed increased in vitro or in vivo RAS signaling pathway activity because of gain-of-function RRAS2 variants. Clinical features are similar to those previously reported, suggesting that RRAS2 gain-of-function variants cause this disease in patients

Emergence of IncX4 plasmids encoding mcr-1 in a clinical isolate of Klebsiella pneumoniae in Japan

The mcr-1 gene encodes a phosphoethanolamine transferase that confers resistance to colistin by transferring phosphoethanolamine to lipid A. A 33-kb IncX4 plasmid harboring mcr-1 was detected in a Klebsiella pneumoniae isolate and two Escherichia coli isolates, and a 66-kb IncI2 plasmid was detected in three E. coli isolates in hospitals in Okinawa, Japan. Keywords: Colistin resistance, MCR-1 producing Klebsiella pneumoniae, IncX4 plasmid harboring mcr-

The genome of an intracellur symbiont of deep sea clam, Calyptogena phaseoliformis, is still in an early phase of reductive genome evolution.

http://www.godac.jamstec.go.jp/darwin/cruise/yokosuka/yk09-12/ehttp://www.godac.jamstec.go.jp/darwin/cruise/yokosuka/yk11-04/

A Carbapenem-Resistant Pseudomonas aeruginosa Isolate Harboring Two Copies of blaIMP-34 Encoding a Metallo-β-Lactamase.

A carbapenem-resistant strain of Pseudomonas aeruginosa, NCGM1984, was isolated in 2012 from a hospitalized patient in Japan. Immunochromatographic assay showed that the isolate was positive for IMP-type metallo-β-lactamase. Complete genome sequencing revealed that NCGM1984 harbored two copies of blaIMP-34, located at different sites on the chromosome. Each blaIMP-34 was present in the same structures of the class 1 integrons, tnpA(ISPa7)-intI1-qacG-blaIMP-34-aac(6')-Ib-qacEdelta1-sul1-orf5-tniBdelta-tniA. The isolate belonged to multilocus sequence typing ST235, one of the international high-risk clones. IMP-34, with an amino acid substitution (Glu126Gly) compared with IMP-1, hydrolyzed all β-lactamases tested except aztreonam, and its catalytic activities were similar to IMP-1. This is the first report of a clinical isolate of an IMP-34-producing P. aeruginosa harboring two copies of blaIMP-34 on its chromosome

Ubiquitin-like 3 as a new protein-sorting factor for small extracellular vesicles

Ubiquitin-like 3 (UBL3) is a well-conserved ubiquitin-like protein (UBL) in eukaryotes and regulates the ubiquitin cascade, but the significant roles of UBL3 in cellular processes remained unknown. Recently, UBL3 was elucidated to be a post-translational modification factor that promotes protein sorting to small extracellular vesicles (sEVs). Proteins sorted into sEVs have been studied as etiologies of sEV-related diseases. Also, there have been attempts to construct drug delivery systems (DDSs) by loading proteins into sEVs. In this review, we introduce the new concept that UBL3 has a critical role in the protein-sorting system and compare structure conservation between UBL3 and other UBLs from an evolutionary perspective. We conclude with future perspectives for the utility of UBL3 in sEV-related diseases and DDS. Key words: UBL3, small extracellular vesicles, protein sorting, ubiquitin-like protein, post-translational modificatio

MICs of β-lactams for <i>P</i>. <i>aeruginosa</i> NCGM1984 and <i>E</i>. <i>coli</i> transformants with <i>bla</i>_IMP-1 and <i>bla</i>_IMP-34.

<p>MICs of β-lactams for <i>P</i>. <i>aeruginosa</i> NCGM1984 and <i>E</i>. <i>coli</i> transformants with <i>bla</i>_IMP-1 and <i>bla</i>_IMP-34.</p