The Rac-GAP alpha2-Chimaerin Signals via CRMP2 and Stathmins in the Development of the Ocular Motor System.

A precise sequence of axon guidance events is required for the development of the ocular motor system. Three cranial nerves grow toward, and connect with, six extraocular muscles in a stereotyped pattern, to control eye movements. The signaling protein alpha2-chimaerin (α2-CHN) plays a pivotal role in the formation of the ocular motor system; mutations in CHN1, encoding α2-CHN, cause the human eye movement disorder Duane Retraction Syndrome (DRS). Our research has demonstrated that the manipulation of α2-chn signaling in the zebrafish embryo leads to ocular motor axon wiring defects, although the signaling cascades regulated by α2-chn remain poorly understood. Here, we demonstrate that several cytoskeletal regulatory proteins-collapsin response mediator protein 2 (CRMP2; encoded by the gene dpysl2), stathmin1, and stathmin 2-bind to α2-CHN. dpysl2, stathmin1, and especially stathmin2 are expressed by ocular motor neurons. We find that the manipulation of dpysl2 and of stathmins in zebrafish larvae leads to defects in both the axon wiring of the ocular motor system and the optokinetic reflex, impairing horizontal eye movements. Knockdowns of these molecules in zebrafish larvae of either sex caused axon guidance phenotypes that included defasciculation and ectopic branching; in some cases, these phenotypes were reminiscent of DRS. chn1 knock-down phenotypes were rescued by the overexpression of CRMP2 and STMN1, suggesting that these proteins act in the same signaling pathway. These findings suggest that CRMP2 and stathmins signal downstream of α2-CHN to orchestrate ocular motor axon guidance and to control eye movements.SIGNIFICANCE STATEMENT The precise control of eye movements is crucial for the life of vertebrate animals, including humans. In humans, this control depends on the arrangement of nerve wiring of the ocular motor system, composed of three nerves and six muscles, a system that is conserved across vertebrate phyla. Mutations in the protein alpha2-chimaerin have previously been shown to cause eye movement disorders (squint) and axon wiring defects in humans. Our recent work has unraveled how alpha2-chimaerin coordinates axon guidance of the ocular motor system in animal models. In this article, we demonstrate key roles for the proteins CRMP2 and stathmin 1/2 in the signaling pathway orchestrated by alpha2-chimaerin, potentially giving insight into the etiology of eye movement disorders in humans.

Engineering a Novel Bivalent Oral Vaccine against Enteric Fever.

Enteric fever is a major global healthcare issue caused largely by Salmonella enterica serovars Typhi and Paratyphi A. The objective of this study was to develop a novel, bivalent oral vaccine capable of protecting against both serovars. Our approach centred on genetically engineering the attenuated S. Typhi ZH9 strain, which has an excellent safety record in clinical trials, to introduce two S. Paratyphi A immunogenic elements: flagellin H:a and lipopolysaccharide (LPS) O:2. We first replaced the native S. Typhi fliC gene encoding flagellin with the highly homologous fliC gene from S. Paratyphi A using Xer-cise technology. Next, we replaced the S. Typhi rfbE gene encoding tyvelose epimerase with a spacer sequence to enable the sustained expression of O:2 LPS and prevent its conversion to O:9 through tyvelose epimerase activity. The resulting new strain, ZH9PA, incorporated these two genetic changes and exhibited comparable growth kinetics to the parental ZH9 strain. A formulation containing both ZH9 and ZH9PA strains together constitutes a new bivalent vaccine candidate that targets both S. Typhi and S. Paratyphi A antigens to address a major global healthcare gap for enteric fever prophylaxis. This vaccine is now being tested in a Phase I clinical trial (NCT04349553).

Inhibition of the diclofenac-induced cyclooxygenase-2 activity by paracetamol in cultured macrophages is not related to the intracellular lipid hydroperoxide tone.

Paracetamol, a weak inhibitor of cyclooxygenase COX-1 and COX-2 activities, has been reported to inhibit the activity of COX-2 induced by diclofenac in J774.2 macrophage cell line. The lack of inhibition of COX-2 by paracetamol in inflamed tissues and thereby the lack of anti-inflammatory activity has been attributed to high lipid hydroperoxide (LHP) tone. In this study, we demonstrate that the inhibition of the diclofenac-induced COX-2 activity in J774.2 cells by paracetamol is not related to the intracellular LHP tone as paracetamol inhibited this activity in the absence and presence of T-butyl hydroperoxide, which is an LHP donor, to the same extents. In fact, treatment of the cells with diclofenac resulted in an increase in the LHP tone. Stimulation of the cells with lipopolysaccharide (LPS) results in the induction of a COX-2 activity, which was not inhibited by paracetamol. This represents the classical induction pathway for COX-2. LPS stimulation did not alter the LHP tone. These results suggest that the enzymatic activity of the diclofenac-induced COX-2 protein does not depend on the supply of hydroperoxides to its peroxidase active site.