Kinesin-1 captures RNA cargo in its adaptable coils

The prototypic and ubiquitous microtubule motor, kinesin-1, uses a variety of adaptor proteins to facilitate the selective transport of diverse cargo within the cell. These cargo adaptors bind to the motor complex through interactions with the kinesin light or heavy chains (KLCs or KHCs). In this issue of Genes & Development, Dimitrova-Paternoga et al. (pp. 976–991) present the first structural characterization of a KHC–cargo adaptor interface. They describe an antiparallel heterotrimeric coiled-coil complex between the carboxy tail of KHC and Tm1-I/C (aTm1), the atypical tropomyosin that is important for oskar mRNA transport in Drosophila oocytes. This interaction enhances direct binding between KHC and RNA. Their findings demonstrate the structural plasticity of the KHC tail as a platform for protein–protein interactions and reveal how a cargo adaptor protein can modify a motor–RNA interface to promote transport

Sarcopenia in Children With End-Stage Liver Disease on the Transplant Waiting List

Sarcopenia predicts morbidity and mortality in adults with end-stage liver disease (ESLD) and is determined by total psoas muscle area (tPMA) measurement from computed tomography (CT) imaging. Recently developed pediatric age- and sex-specific tPMA growth curves provide the opportunity to ascertain prevalence and impact of sarcopenia in children awaiting liver transplantation (LT). This retrospective single-center study evaluated sarcopenia in children between 1 and 16 years with ESLD and a clinically indicated abdominal CT less than 3 months before first isolated LT. Sarcopenia was defined as tPMA z score less than −2 measured at the intervertebral L4-5 level. Patient demographic, biochemical, and outcome data were recorded. tPMA was compared with other measures of nutritional status using univariate and multivariate logistic analyses. Outcome measures included 1-year morbidity events and mortality after LT. CT images from 25 (64% female) children with median age of 5.50 (interquartile range [IQR], 3.75-11.33) years were reviewed. Ten children (40%) had a tPMA z score less than −2. Sarcopenia was associated with lower z scores for weight (odds ratio [OR], 0.38; P = 0.02), height (OR, 0.32; P = 0.03), and nutritional support before LT (OR, 12.93; P = 0.01). Sarcopenic children had a longer duration of pediatric intensive care unit (PICU) stay (3.50 [IQR, 3.00-6.00] versus 2.00 [IQR, 2.00-3.50] days; P = 0.03). Sarcopenia was prevalent in 40% of children with ESLD awaiting LT, and lower tPMA z score was associated with deficient anthropometrics and need for nutritional support before LT. Post-LT PICU duration was increased in children with sarcopenia, reflecting adverse outcomes associated with muscle loss. Further studies are needed to elucidate the underlying mechanisms of sarcopenia in children with ESLD

Paediatric reference values for total psoas muscle area

Background: Sarcopenia, the unintentional loss of skeletal muscle mass, is associated with poor outcomes in adult patient populations. In adults, sarcopenia is often ascertained by cross-sectional imaging of the psoas muscle area (PMA). Although children with chronic medical illnesses may be at increased risk for muscle loss because of nutritional deficiencies, physical deconditioning, endocrine anomalies, and systemic inflammation, consistent quantitative definitions for sarcopenia in children are lacking. We aimed to generate paediatric reference values for PMA at two intervertebral lumbar levels, L3–4 and L4–5. Methods: In this cross-sectional study, we analysed abdominal computed tomography scans of consecutive children presenting to the emergency department. Participants were children 1–16 years who required abdominal cross-sectional imaging after paediatric trauma between January 1, 2005 and December 31, 2015 in a large Canadian quaternary care centre. Children with a documented chronic medical illness or an acute spinal trauma at presentation were excluded. Total PMA (tPMA) at levels L3–4 and L4–5 were measured in square millimetres (mm2) as the sum of left and right PMA. Age-specific and sex-specific tPMA percentile curves were modelled using quantile regression. Results: Computed tomography images from 779 children were included. Values of tPMA at L4–5 were significantly larger than at L3–4 at all ages, but their correlation was high for both girls (r = 0.95) and boys (r = 0.98). Amongst girls, tPMA 50th percentile values ranged from 365 to 2336 mm2 at L3–4 and from 447 to 2704 mm2 for L4–5. Amongst boys, 50th percentile values for tPMA ranged between 394 and 3050 mm2 at L3–4 and from 498 to 3513 mm2 at L4–5. Intraclass correlation coefficients were excellent at L3–4 (0.97, 95% CI 0.94 to 0.981) and L4–5 (0.99, 95% CI 0.986 to 0.995). Weight and tPMA were correlated, stratified by sex for boys (L3–4 r = 0.90; L4–5 r = 0.90) and for girls (L3–4 r = 0.87; L4–5 r = 0.87). An online application was subsequently developed to easily calculate age-specific and sex-specific z-scores and percentiles. Conclusions: We provide novel paediatric age-specific and sex-specific growth curves for tPMA at intervertebral L3–4 and L4–5 levels for children between the ages of 1-16 years. Together with an online tool (https://ahrc-apps.shinyapps.io/sarcopenia/), these tPMA curves should serve as a reference enabling earlier identification and targeted intervention of sarcopenia in children with chronic medical conditions

Molecular mechanism for kinesin-1 direct membrane recognition

The cargo-binding capabilities of cytoskeletal motor proteins have expanded during evolution through both gene duplication and alternative splicing. For the light chains of the kinesin-1 family of microtubule motors, this has resulted in an array of carboxyl-terminal domain sequences of unknown molecular function. Here, combining phylogenetic analyses with biophysical, biochemical, and cell biology approaches, we identify a highly conserved membrane-induced curvature-sensitive amphipathic helix within this region of a subset of long kinesin light-chain paralogs and splice isoforms. This helix mediates the direct binding of kinesin-1 to lipid membranes. Membrane binding requires specific anionic phospholipids, and it contributes to kinesin-1\u2013dependent lysosome positioning, a canonical activity that, until now, has been attributed exclusively the recognition of organelle-associated cargo adaptor proteins. This leads us to propose a protein-lipid coincidence detection framework for kinesin-1\u2013mediated organelle transport

Fragment-linking peptide design yields a high-affinity ligand for microtubule-based transport

Synthetic peptides are attractive candidates to manipulate protein-protein interactions inside the cell as they mimic natural interactions to compete for binding. However, protein-peptide interactions are often dynamic and weak. A challenge is to design peptides that make improved interactions with the target. Here, we devise a fragment-linking strategy\u2014\u201cmash-up\u201d design\u2014to deliver a high-affinity ligand, KinTag, for the kinesin-1 motor. Using structural insights from natural micromolar-affinity cargo-adaptor ligands, we have identified and combined key binding features in a single, high-affinity ligand. An X-ray crystal structure demonstrates interactions as designed and reveals only a modest increase in interface area. Moreover, when genetically encoded, KinTag promotes transport of lysosomes with higher efficiency than natural sequences, revealing a direct link between motor-adaptor binding affinity and organelle transport. Together, these data demonstrate a fragment-linking strategy for peptide design and its application in a synthetic motor ligand to direct cellular cargo transport

Conservative management of an acquired tracheoesophageal fistula caused by a swallowed button battery: A case report

Introduction: The incidence of pediatric button battery (BB) ingestions has increased in recent years with a subsequent rise in secondary esophageal injuries. There is a paucity of published literature on the evaluation and management of acquired trachea-esophageal fistulas (TEF) in this context and therefore, the purpose of this paper was to summarize a case and propose an evidence-based protocol for acquired TEF management. Case presentation: A 15-month-old female presenting with fever, mild tachypnea and lethargy underwent a chest x-ray which demonstrated a BB in the upper thorax. Laryngoscopy, and subsequent rigid esophagoscopy, identified the BB in the proximal esophagus. There was surrounding inflammation and liquefactive necrosis, and the BB was removed via rigid esophagoscopy. A post-procedure chest computerized tomography following BB removal revealed a small collection in the mediastinum consistent with full thickness esophageal perforation as well as mediastinitis. She was treated conservatively and on post-operative day (POD) 7, a swallow study demonstrated no evidence of leak or stricture, thus a trial of oral, clear fluids took place. A next-day repeat swallow study was performed and revealed an acquired TEF. She was treated conservatively with proton pump inhibitor and nasojejunal feeds. Bronchoscopy, upper gastrointestinal endoscopy and repeat esophagram at six-weeks demonstrated resolution of the TEF. Conclusion: Clinicians should have a high index of suspicion for BB ingestion complications, even after removal. This case illustrated that patients with acquired TEF can be managed conservatively. Following literature review and our personal experience in this single case, we propose a suggested algorithm for management of these patients

De novo designed peptides for cellular delivery and subcellular localisation

Increasingly, it is possible to design peptide and protein assemblies de novo from first principles or computationally. This approach provides new routes to functional synthetic polypeptides, including designs to target and bind proteins of interest. Much of this work has been developed in vitro. Therefore, a challenge is to deliver de novo polypeptides efficiently to sites of action within cells. Here we describe the design, characterisation, intracellular delivery, and subcellular localisation of a de novo synthetic peptide system. This system comprises a dual-function basic peptide, programmed both for cell penetration and target binding, and a complementary acidic peptide that can be fused to proteins of interest and introduced into cells using synthetic DNA. The designs are characterised in vitro using biophysical methods and X-ray crystallography. The utility of the system for delivery into mammalian cells and subcellular targeting is demonstrated by marking organelles and actively engaging functional protein complexes

Molecular architecture of the autoinhibited kinesin-1 lambda particle

Despite continuing progress in kinesin enzyme mechanochemistry and emerging understanding of the cargo recognition machinery, it is not known how these functions are coupled and controlled by the α-helical coiled coils encoded by a large component of kinesin protein sequences. Here, we combine computational structure prediction with single-particle negative-stain electron microscopy to reveal the coiled-coil architecture of heterotetrameric kinesin-1 in its compact state. An unusual flexion in the scaffold enables folding of the complex, bringing the kinesin heavy chain–light chain interface into close apposition with a tetrameric assembly formed from the region of the molecule previously assumed to be the folding hinge. This framework for autoinhibition is required to uncover how engagement of cargo and other regulatory factors drives kinesin-1 activation