Hydrophobic Transmembrane Domain Research Articles

Carbocation Footprinting of Soluble and Transmembrane Proteins

Here, we introduce carbocations (R3C+) as laser-initiated footprinting reagents for proteins. We screened seven candidates and selected trifluomethoxy benzyl bromide (TFBB) as an effective precursor for the electrophilic trifluomethoxy benzyl carbocation (TFB+) under laser (248 nm) irradiation on the fast photochemical oxidation of proteins (FPOP) platform. Initial results demonstrate that this electrophilic cation reagent affords residue coverage of nucleophilic amino acids including H, W, M, and S. Further, the addition of TFB+ increases the hydrophobicity of the peptides so that separation of isomeric peptide products by reversed-phase LC is improved, suggesting opportunities for subresidue footprinting. Comparison of apo- and holo-myoglobin footprints shows that the TFB+ footprinting is sensitive to protein conformational change and solvent accessibility. Interestingly, because the TFB+ is amphiphilic, the reagent can potentially footprint membrane proteins as demonstrated for vitamin K epoxide reductase (VKOR) stabilized in a micelle. Not only does footprinting of the extra-membrane domain occur, but also some footprinting of the hydrophobic transmembrane domain is achieved owing to the interaction of TFB+ with the micelle. Carbocation precursors are stable and amenable for tailoring their properties and those of the incipient carbocation, enabling targeting their soluble or membrane-associated or embedded regions and distinguishing between the extra- and trans-membrane domains of membrane proteins.

Sequence-based features that are determinant for tail-anchored membrane protein sorting in eukaryotes.

The correct targeting and insertion of tail-anchored (TA) integral membrane proteins is critical for cellular homeostasis. TA proteins are defined by a hydrophobic transmembrane domain (TMD) at their C-terminus and are targeted to either the ER or mitochondria. Derived from experimental measurements of a few TA proteins, there has been little examination of the TMD features that determine localization. As a result, the localization of many TA proteins are misclassified by the simple heuristic of overall hydrophobicity. Because ER-directed TMDs favor arrangement of hydrophobic residues to one side, we sought to explore the role of geometric hydrophobic properties. By curating TA proteins with experimentally determined localizations and assessing hypotheses for recognition, we bioinformatically and experimentally verify that a hydrophobic face is the most accurate singular metric for separating ER and mitochondria-destined yeast TA proteins. A metric focusing on an 11 residue segment of the TMD performs well when classifying human TA proteins. The most inclusive predictor uses both hydrophobicity and C-terminal charge in tandem. This work provides context for previous observations and opens the door for more detailed mechanistic experiments to determine the molecular factors driving this recognition.

LPXRFa and its receptor in yellowtail kingfish (Seriola lalandi): Molecular cloning, ontogenetic expression profiles, and stimulatory effects on growth hormone and gonadotropin gene expression

Despite its functional significance in mammals and birds, the biological role of gonadotropin-inhibitory hormone (GnIH) in reproduction is still far from being fully understood in teleosts. In the current study, we have identified LPXRFa, the piscine ortholog of GnIH, and its cognate receptor (LPXRFa-R) in yellowtail kingfish (YTK), which is considered as a promising species for aquaculture industry worldwide. The YTK cDNA sequence of lpxrfa was 534 base pair (bp) in length and encoded a 178-amino acids (aa) preprohormone. The LPXRFa precursor comprised three putative peptide sequences that included –MPMRF, –MPQRF, or -LPERL motifs at the C-termini, respectively. The YTK lpxrfa-r cDNA sequence was composed of 1265 bp that gave rise to a LPXRFa-R of 420 aa, encompassing the characteristic seven hydrophobic transmembrane domains. In males, both lpxrfa and lpxrfa-r transcripts could be detected at high levels in the brain and testis. In females, a noteworthy expression of lpxrfa was observed in the brain and ovary, while the expression of lpxrfa-r was especially evident only in the brain. To study the ontogeny of LPXRFa system, transcript levels were also investigated during early life stages. Variable expression of the LPXRFa system was observed during all stages of YTK embryogenesis. The highest expression of lpxrfa and lpxrfa-r were noticed at 7 dph and 15 dph, respectively. Furthermore, LPXRFa peptides stimulated growth hormone (gh), luteinizing hormone (lhβ) and follicle-stimulating hormone (fshβ) gene expression from the pituitary. Taken together, our results provide initial evidence for the existence of the LPXRFa system in yellowtail kingfish and suggest its possible involvement at early development and reproductive functions.

The role of the SARS-CoV-2 envelope protein as a pH-dependent cation channel.

The role of the SARS-CoV-2 envelope protein as a pH-dependent cation channel.

Looking for a safe haven: tail-anchored proteins and their membrane insertion pathways.

Insertion of membrane proteins into the lipid bilayer is a crucial step during their biosynthesis. Eukaryotic cells face many challenges in directing these proteins to their predestined target membrane. The hydrophobic signal peptide or transmembrane domain (TMD) of the nascent protein must be shielded from the aqueous cytosol and its target membrane identified followed by transport and insertion. Components that evolved to deal with each of these challenging steps range from chaperones to receptors, insertases, and sophisticated translocation complexes. One prominent translocation pathway for most proteins is the signal recognition particle (SRP)-dependent pathway which mediates co-translational translocation of proteins across or into the endoplasmic reticulum (ER) membrane. This textbook example of protein insertion is stretched to its limits when faced with secretory or membrane proteins that lack an amino-terminal signal sequence or TMD. Particularly, a large group of so-called tail-anchored (TA) proteins that harbor a single carboxy-terminal TMD require an alternative, post-translational insertion route into the ER membrane. In this review, we summarize the current research in TA protein insertion with a special focus on plants, address challenges, and highlight future research avenues.

Tetraspanins: Novel Molecular Regulators of Gastric Cancer.

Gastric cancer is the fourth and fifth most common cancer worldwide in men and women, respectively. However, patients with an advanced stage of gastric cancer still have a poor prognosis and low overall survival rate. The tetraspanins belong to a protein superfamily with four hydrophobic transmembrane domains and 33 mammalian tetraspanins are ubiquitously distributed in various cells and tissues. They interact with other membrane proteins to form tetraspanin-enriched microdomains and serve a variety of functions including cell adhesion, invasion, motility, cell fusion, virus infection, and signal transduction. In this review, we summarize multiple utilities of tetraspanins in the progression of gastric cancer and the underlying molecular mechanisms. In general, the expression of TSPAN8, CD151, TSPAN1, and TSPAN4 is increased in gastric cancer tissues and enhance the proliferation and invasion of gastric cancer cells, while CD81, CD82, TSPAN5, TSPAN9, and TSPAN21 are downregulated and suppress gastric cancer cell growth. In terms of cell motility regulation, CD9, CD63 and CD82 are metastasis suppressors and the expression level is inversely associated with lymph node metastasis. We also review the clinicopathological significance of tetraspanins in gastric cancer including therapeutic targets, the development of drug resistance and prognosis prediction. Finally, we discuss the potential clinical value and current limitations of tetraspanins in gastric cancer treatments, and provide some guidance for future research.

Deubiquitinases USP20/33 promote the biogenesis of tail-anchored membrane proteins.

Numerous proteins that have hydrophobic transmembrane domains (TMDs) traverse the cytosol and posttranslationally insert into cellular membranes. It is unclear how these hydrophobic membrane proteins evade recognition by the cytosolic protein quality control (PQC), which typically recognizes exposed hydrophobicity in misfolded proteins and marks them for proteasomal degradation by adding ubiquitin chains. Here, we find that tail-anchored (TA) proteins, a vital class of membrane proteins, are recognized by cytosolic PQC and are ubiquitinated as soon as they are synthesized in cells. Surprisingly, the ubiquitinated TA proteins are not routed for proteasomal degradation but instead are handed over to the targeting factor, TRC40, and delivered to the ER for insertion. The ER-associated deubiquitinases, USP20 and USP33, remove ubiquitin chains from TA proteins after their insertion into the ER. Thus, our data suggest that deubiquitinases rescue posttranslationally targeted membrane proteins that are inappropriately ubiquitinated by PQC in the cytosol.

Molecular basis of tail-anchored integral membrane protein recognition by the cochaperone Sgt2

The targeting and insertion of tail-anchored (TA) integral membrane proteins (IMPs) into the correct membrane is critical for cellular homeostasis. The fungal protein Sgt2, and its human homolog SGTA, is the entry point for clients to the guided entry of tail-anchored protein (GET) pathway, which targets endoplasmic reticulum-bound TA IMPs. Consisting of three structurally independent domains, the C terminus of Sgt2 binds to the hydrophobic transmembrane domain (TMD) of clients. However, the exact binding interface within Sgt2 and molecular details that underlie its binding mechanism and client preference are not known. Here, we reveal the mechanism of Sgt2 binding to hydrophobic clients, including TA IMPs. Through sequence analysis, biophysical characterization, and a series of capture assays, we establish that the Sgt2 C-terminal domain is flexible but conserved and sufficient for client binding. A molecular model for this domain reveals a helical hand forming a hydrophobic groove approximately 15 Å long that is consistent with our observed higher affinity for client TMDs with a hydrophobic face and a minimal length of 11 residues. This work places Sgt2 into a broader family of TPR-containing cochaperone proteins, demonstrating structural and sequence-based similarities to the DP domains in the yeast Hsp90 and Hsp70 coordinating protein, Sti1.

Influenza A H1 and H3 Transmembrane Domains Interact Differently with Each Other and with Surrounding Membrane Lipids.

Hemagglutinin (HA) is a class I viral membrane fusion protein, which is the most abundant transmembrane protein on the surface of influenza A virus (IAV) particles. HA plays a crucial role in the recognition of the host cell, fusion of the viral envelope and the host cell membrane, and is the major antigen in the immune response during the infection. Mature HA organizes in homotrimers consisting of a sequentially highly variable globular head and a relatively conserved stalk region. Every HA monomer comprises a hydrophilic ectodomain, a pre-transmembrane domain (pre-TMD), a hydrophobic transmembrane domain (TMD), and a cytoplasmic tail (CT). In recent years the effect of the pre-TMD and TMD on the structure and function of HA has drawn some attention. Using bioinformatic tools we analyzed all available full-length amino acid sequences of HA from 16 subtypes across various host species. We calculated several physico-chemical parameters of HA pre-TMDs and TMDs including accessible surface area (ASA), average hydrophobicity (Hav), and the hydrophobic moment (µH). Our data suggests that distinct differences in these parameters between the two major phylogenetic groups, represented by H1 and H3 subtypes, could have profound effects on protein–lipid interactions, trimer formation, and the overall HA ectodomain orientation and antigen exposure.

Structural insights into the membrane receptor ShuA in DDM micelles and in a model of gram-negative bacteria outer membrane as seen by SAXS and MD simulations

Successful crystallization of membrane proteins in detergent micelles depends on key factors such as conformational stability of the protein in micellar assemblies, the protein-detergent complex (PDC) monodispersity and favorable protein crystal contacts by suitable shielding of the protein hydrophobic surface by the detergent belt. With the aim of studying the influence of amphiphilic environment on membrane protein structure, stability and crystallizability, we combine molecular dynamics (MD) simulations with SEC-MALLS and SEC-SAXS (Size Exclusion Chromatography in line with Multi Angle Laser Light Scattering or Small Angle X-ray Scattering) experiments to describe the protein-detergent interactions that could help to rationalize PDC crystallization. In this context, we compare the protein-detergent interactions of ShuA from Shigella dysenteriae in n-Dodecyl-β-D-Maltopyranoside (DDM) with ShuA inserted in a realistic model of gram-negative bacteria outer membrane (OM) containing a mixture of bacterial lipopolysaccharide and phospholipids. To evaluate the quality of the PDC models, we compute the corresponding SAXS curves from the MD trajectories and compare with the experimental ones. We show that computed SAXS curves obtained from the MD trajectories reproduce better the SAXS obtained from the SEC-SAXS experiments for ShuA surrounded by 268 DDM molecules. The MD results show that the DDM molecules form around ShuA a closed belt whose the hydrophobic thickness appears slightly smaller (~22 Å) than the hydrophobic transmembrane domain of the protein (24.6 Å) suggested by Orientations of Proteins in Membranes (OPM) database. The simulations also show that ShuA transmembrane domain is remarkably stable in all the systems except for the extracellular and periplasmic loops that exhibit larger movements due to specific molecular interactions with lipopolysaccharides (LPS). We finally point out that this detergent behavior may lead to the occlusion of the periplasmic hydrophilic surface and poor crystal contacts leading to difficulties in crystallization of ShuA in DDM.

The roles of tetraspanins in bacterial infections.

Tetraspanins, a wide family composed of 33 transmembrane proteins, are associated with different types of proteins through which they arbitrate important cellular processes such as fusion, adhesion, invasion, tissue differentiation and immunological responses. Tetraspanins share a comparable structural design, which consists of four hydrophobic transmembrane domains with cytoplasmic and extracellular loops. They cooperate with different proteins, including other tetraspanins, receptors or signalling proteins to compose functional complexes at the cell surface, designated tetraspanin-enriched microdomains (TEM). Increasing evidences establish that tetraspanins are exploited by numerous intracellular pathogens as a doorway for entering and replicating within human cells. Although previous surveys focused mainly on viruses and parasites, it is now becoming clear that bacteria interact with tetraspanins, using TEM as a "gateway" to infection. In this review, we examine the biological functions of tetraspanins that are relevant to bacterial infective procedures and consider the available data that reveal how different bacteria benefit from host cell tetraspanins in infection and in the pathogenesis of diseases. We will also emphasise the stimulating potentials of targeting tetraspanins for preventing bacterial infectious diseases, using specific neutralising antibodies or anti-adhesion peptide-based therapies. Such innovative therapeutic opportunities may deliver alternatives for fighting difficult-to-manage and drug-resistant bacterial pathogens.

Immunological regulation by a β-adrenergic-like octopamine receptor gene in crowded larvae of the oriental Armyworm, Mythmina separata

Recent reports demonstrate that octopamine plays an important immunological role in crowded larvae of the Oriental Armyworm, Mythmina separata. We identified an octopamine receptor, the β-adrenergic-like gene (designated MsOctβ2R), with a 1191 bp open reading frame that encodes 396 amino acids and contains seven conserved hydrophobic transmembrane domains. Multiple sequence alignments and a phylogenetic analysis indicated that MsOctβ2R was orthologous to Octβ2R that is present in other lepidopterans. MsOctβ2R was expressed throughout all developmental stages with higher relative expression during the fourth instar and adult stages. MsOctβ2R was highly expressed in the ventral nerve cord and the fat body relative to other examined tissues. Elevated MsOctβ2R expression was observed in larvae that were under higher-density conditions (7 and 10 larvae per jar). Silencing MsOctβ2R expression via dsRNA injections in larvae from higher-density conditions significantly decreased phenoloxidase (PO) and lysozyme activity, total haemocyte counts, and survival rates against Beauveria bassiana infections (54.06%, 9.91%, 36.22%, and 23.53%, respectively) when compared with control larvae. These results suggest that high-density conditions might alter prophylactic immunity in larvae by regulating the MsOctβ2R gene in M. separara and provide new insights into density-dependent prophylaxis in insects.

Ayu (Plecoglossus altivelis) CD46 isoforms protect the cells from autologous complement attack

CD46 is an important immune regulatory receptor with multiple functions. However, studies on the function of teleost CD46, especially the different CD46 isoforms are limited. In this study, we identified three membrane cofactor protein (MCP, CD46) gene isoforms from ayu (Plecoglossus altivelis) and tentatively named as PaCD46 isoforms. PaCD46 isoforms were generated by alternative splicing and all consisted of four conserved short consensus repeats (SCRs), and the variable serine-threonine-proline-rich domain, transmembrane hydrophobic domain, and cytoplasmic tail. Phylogenetic analysis showed that the isoforms clustered together with other fish CD46 and then with higher animal CD46. Western blotting analysis of peripheral blood mononuclear cells (PBMC) revealed three bands, all of which had much larger molecular weights than the theoretical values of the three PaCD46 isoforms. Moreover, three PaCD46 isoforms were individually expressed on HEK293 cells, and Western blotting showed the similar band profile to that of PBMC. The recombinant extracellular domain of the PaCD46 isoforms, obtained by expression in Pichia pastoris, significantly reduced hemolysis activity of ayu sera. Furthermore, each of the three PaCD46 isoforms respectively protected the HEK293 cells expressing the isoform. The isoforms were also identified for their protection of autologous PBMC from complement activation. These results provided the first evidence that PaCD46 isoforms may be complement regulatory proteins to prevent complement-induced damage to self-tissue.

Differential Modes of Orphan Subunit Recognition for the WRB/CAML Complex

SUMMARYA large proportion of membrane proteins must be assembled into oligomeric complexes for function. How this process occurs is poorly understood, but it is clear that complex assembly must be tightly regulated to avoid accumulation of orphan subunits with potential cytotoxic effects. We interrogated assembly in mammalian cells by using the WRB/CAML complex, an essential insertase for tail-anchored proteins in the endoplasmic reticulum (ER), as a model system. Our data suggest that the stability of each subunit is differentially regulated. In WRB’s absence, CAML folds incorrectly, causing aberrant exposure of a hydrophobic transmembrane domain to the ER lumen. When present, WRB can correct the topology of CAML both in vitro and in cells. In contrast, WRB can independently fold correctly but is still degraded in the absence of CAML. We therefore propose that there are at least two distinct regulatory pathways for the surveillance of orphan subunits in the mammalian ER.

Identification, characterization, and expression analysis of a serotonin receptor involved in the reproductive process of the Pacific abalone, Haliotis discus hannai.

Serotonin receptor (5-HT) is a biogenic amine acting as a neurotransmitter and neuromodulator that mediates various aspects of reproduction and gametogenesis. The full-length nucleotide sequence of Haliotis discus hannai encodes a protein of 417 amino acids with a predicted molecular mass of 46.54kDa and isoelectric point of 8.94. The structural profile of 5-HTHdh displayed key features of G protein-coupled receptors, including seven hydrophobic transmembrane domains, putative N-linked glycosylation sites, and several phosphorylation consensus motifs. It shares the highest homology of its amino acid sequence with the 5-HT receptor from Haliotis asinina, and to lesser extent of human 5-HT receptor. The cloned sequence possesses two cysteine residues (Cys-115 and Cys-193), which are likely to form a disulfide bond. Phylogenetic comparison with other known 5-HT receptor genes revealed that the 5-HTHdh is most closely related to the 5-HTHa receptor. The three-dimensional structure of the 5-HTHdh showed multiple alpha helices which is separated by a helix-loop-helix (HLH) structure. Quantitative PCR demonstrated that the receptor mRNA was predominantly expressed in the pleuropedal ganglion. Significant differences in the transcriptional activity of the 5-HTHdh gene were observed in the ovary at the ripening stage. An exclusive expression was detected in pleuropedal ganglion, testis, and ovary at higher effective accumulative temperature (1000°C). In situ hybridization showed that the 5-HTHdh expressing neurosecretory cells were distributed in the cortex of the pleuropedal ganglion. Our results suggest that 5-HTHdh synthesized in the neural ganglia may be involved in oocyte maturation and spawning of H. discus hannai.

Distinct Requirements for Tail-Anchored Membrane Protein Biogenesis in Escherichia coli.

Tail-anchored membrane proteins (TAMPs) are a distinct subset of inner membrane proteins (IMPs) characterized by a single C-terminal transmembrane domain (TMD) that is responsible for both targeting and anchoring. Little is known about the routing of TAMPs in bacteria. Here, we have investigated the role of TMD hydrophobicity in tail-anchor function in Escherichia coli and its influence on the choice of targeting/insertion pathway. We created a set of synthetic, fluorescent TAMPs that vary in the hydrophobicity of their TMDs and corresponding control polypeptides that are extended at their C terminus to create regular type II IMPs. Surprisingly, we observed that TAMPs have a much lower TMD hydrophobicity threshold for efficient targeting and membrane insertion than their type II counterparts. Using strains conditional for the expression of known membrane-targeting and insertion factors, we show that TAMPs with strongly hydrophobic TMDs require the signal recognition particle (SRP) for targeting. Neither the SecYEG translocon nor YidC appears to be essential for the membrane insertion of any of the TAMPs studied. In contrast, corresponding type II IMPs with a TMD of sufficient hydrophobicity to promote membrane insertion followed an SRP- and SecYEG translocon-dependent pathway. Together, these data indicate that the capacity of a TMD to promote the biogenesis of E. coli IMPs is strongly dependent upon the polypeptide context in which it is presented.IMPORTANCE A subset of membrane proteins is targeted to and inserted into the membrane via a hydrophobic transmembrane domain (TMD) that is positioned at the very C terminus of the protein. The biogenesis of these so-called tail-anchored proteins (TAMPs) has been studied in detail in eukaryotic cells. Various partly redundant pathways were identified, the choice for which depends in part on the hydrophobicity of the TMD. Much less is known about bacterial TAMPs. The significance of our research is in identifying the role of TMD hydrophobicity in the routing of E. coli TAMPs. Our data suggest that both the nature of the TMD and its role in routing can be very different for TAMPs versus "regular" membrane proteins. Elucidating these position-specific effects of TMDs will increase our understanding of how prokaryotic cells face the challenge of producing a wide variety of membrane proteins.

Identification of a dopamine receptor in Sinonovacula constricta and its antioxidant responses

The dopamine (DA) D2 receptor is a member of the G protein-coupled receptors of organisms and plays an important role in immune system regulation. The presence of DA receptors has been widely reported in vertebrates, but few studies have been conducted in shellfish. Here, we identified a novel DA-D2 receptor gene, ScDopR2-1, in the razor clam Sinonovacula constricta. ScDopR2-1 belongs to the family of G protein-coupled receptors, containing seven hydrophobic transmembrane domains, along with 16 predicted N-glycosylation sites and 69 phosphorylation sites. A longer third intracellular loop and a shorter C-terminus in ScDopR2-1 are characteristic features of D2 receptors. ScDopR2-1 is widely expressed in tissues from adult clams, showing high expression in siphon and foot tissues. Furthermore, in response to Vibrio anguillarum challenge, ScDopR2-1 expression levels are significantly increased in liver tissue. Moreover, changes in the activities of catalase (CAT) and superoxide dismutase (SOD) also indicate that the organism causes an immune response. In summary, the results indicate that ScDopR2-1 plays a pivotal role in antioxidant responses in S. constricta.

Fully hydrophobic HIV gp41 adopts a hemifusion-like conformation in phospholipid bilayers

The HIV envelope glycoprotein mediates virus entry into target cells by fusing the virus lipid envelope with the cell membrane. This process requires large-scale conformational changes of the fusion protein gp41. Current understanding of the mechanisms with which gp41 induces membrane merger is limited by the fact that the hydrophobic N-terminal fusion peptide (FP) and C-terminal transmembrane domain (TMD) of the protein are challenging to characterize structurally in the lipid bilayer. Here we have expressed a gp41 construct that contains both termini, including the FP, the fusion peptide-proximal region (FPPR), the membrane-proximal external region (MPER), and the TMD. These hydrophobic domains are linked together by a shortened water-soluble ectodomain. We reconstituted this "short NC" gp41 into a virus-mimetic lipid membrane and conducted solid-state NMR experiments to probe the membrane-bound conformation and topology of the protein. 13C chemical shifts indicate that the C-terminal MPER-TMD is predominantly α-helical, whereas the N-terminal FP-FPPR exhibits β-sheet character. Water and lipid 1H polarization transfer to the protein revealed that the TMD is well-inserted into the lipid bilayer, whereas the FPPR and MPER are exposed to the membrane surface. Importantly, correlation signals between the FP-FPPR and the MPER are observed, providing evidence that the ectodomain is sufficiently collapsed to bring the N- and C-terminal hydrophobic domains into close proximity. These results support a hemifusion-like model of the short NC gp41 in which the ectodomain forms a partially folded hairpin that places the FPPR and MPER on the opposing surfaces of two lipid membranes.

Insect ATP-Binding Cassette (ABC) Transporters: Roles in Xenobiotic Detoxification and Bt Insecticidal Activity.

ATP-binding cassette (ABC) transporters, a large class of transmembrane proteins, are widely found in organisms and play an important role in the transport of xenobiotics. Insect ABC transporters are involved in insecticide detoxification and Bacillus thuringiensis (Bt) toxin perforation. The complete ABC transporter is composed of two hydrophobic transmembrane domains (TMDs) and two nucleotide binding domains (NBDs). Conformational changes that are needed for their action are mediated by ATP hydrolysis. According to the similarity among their sequences and organization of conserved ATP-binding cassette domains, insect ABC transporters have been divided into eight subfamilies (ABCA–ABCH). This review describes the functions and mechanisms of ABC transporters in insecticide detoxification, plant toxic secondary metabolites transport and insecticidal activity of Bt toxin. With improved understanding of the role and mechanisms of ABC transporter in resistance to insecticides and Bt toxins, we can identify valuable target sites for developing new strategies to control pests and manage resistance and achieve green pest control.

Neurofascin (NFASC) gene mutation causes autosomal recessive ataxia with demyelinating neuropathy

IntroductionNeurofascin, encoded by NFASC, is a transmembrane protein that plays an essential role in nervous system development and node of Ranvier function. Anti-Neurofascin autoantibodies cause a specific type of chronic inflammatory demyelinating polyneuropathy (CIDP) often characterized by cerebellar ataxia and tremor. Recently, homozygous NFASC mutations were recently associated with a neurodevelopmental disorder in two families. MethodsA combined approach of linkage analysis and whole-exome sequencing was performed to find the genetic cause of early-onset cerebellar ataxia and demyelinating neuropathy in two siblings from a consanguineous Italian family. Functional studies were conducted on neurons from induced pluripotent stem cells (iPSCs) generated from the patients. ResultsGenetic analysis revealed a homozygous p.V1122E mutation in NFASC. This mutation, affecting a highly conserved hydrophobic transmembrane domain residue, led to significant loss of Neurofascin protein in the iPSC-derived neurons of affected siblings. ConclusionsThe identification of NFASC mutations paves the way for genetic research in the developing field of nodopathies, an emerging pathological entity involving the nodes of Ranvier, which are associated for the first time with a hereditary ataxia syndrome with neuropathy.