Preventing controlled substance diversion in perioperative settings: a narrative review.

PURPOSE: Diversion of controlled substances in the perioperative setting is an ongoing challenge, with consequences for patients, anesthesiologists, perioperative staff, and health care facilities alike. Perioperative environments are at high risk for diversion, since controlled substances are frequently handled in these settings, with varying levels of oversight. In this narrative review, we summarize strategies for preventing diversion of controlled substances in perioperative settings (i.e., operating rooms, endoscopy suites, and postanesthesia recovery units). SOURCE: We performed a targeted literature search in PubMed MEDLINE, Embase, Scopus, Web of Science, the Cochrane Register of Controlled trials, and the Cochrane Database of Systematic Reviews, as well as a manual search for additional references. We used terminology related to drug diversion, drug abuse, anesthesiologists, pharmacists, physicians, operating room personnel, and controlled substances. PRINCIPAL FINDINGS: Many strategies have been described for preventing diversion in perioperative settings, and these are broadly categorized into: education, distribution, auditing, or provider screening. Some of these approaches may be time- and resource-intensive. There is limited evidence to inform anesthesia departments' choice of which strategies to adopt. CONCLUSION: Although awareness of perioperative controlled substance diversion has been improving, there are too few data to suggest an optimal approach. Anesthesia departments will need to work collaboratively with hospital pharmacies and actively select strategies that are reasonable given local resources.

RéSUMé: OBJECTIF: Le détournement des substances contrôlées en milieu périopératoire constitue un défi permanent qui a des conséquences pour la patientèle, les anesthésiologistes, le personnel périopératoire et les établissements de soins de santé. Les environnements périopératoires courent un risque élevé de détournement, car les substances contrôlées sont fréquemment manipulées dans ces milieux, avec divers niveaux de surveillance. Dans ce compte rendu narratif, nous résumons les stratégies de prévention du détournement des substances contrôlées dans les milieux périopératoires (c.-à-d. salles d'opération, salles d'endoscopie et salles de réveil). SOURCES: Nous avons réalisé une recherche documentaire ciblée dans les bases de données PubMed, MEDLINE, Embase, Scopus, Web of Science, le registre Cochrane des essais contrôlés et la base de données Cochrane des revues systématiques, ainsi qu'une recherche manuelle de références supplémentaires. Nous avons utilisé une terminologie liée au détournement de médicaments, à l'abus de substances, aux anesthésiologistes, aux pharmacien·nes, aux médecins, au personnel de salle d'opération et aux substances contrôlées. CONSTATATIONS PRINCIPALES: De nombreuses stratégies ont été décrites pour prévenir le détournement dans les milieux périopératoires, et celles-ci sont généralement classées en éducation, distribution, audit ou dépistage des fournisseurs et fournisseuses de soin. Certaines de ces approches peuvent exiger beaucoup de temps et de ressources. Il existe peu de données probantes pour éclairer le choix des départements d'anesthésie quant aux stratégies à adopter. CONCLUSION: Bien que la sensibilisation au détournement périopératoire de substances contrôlées se soit améliorée, il y a trop peu de données pour suggérer une approche optimale. Les départements d'anesthésie devront travailler en collaboration avec les pharmacies hospitalières et choisir activement des stratégies raisonnables qui tiennent compte des ressources locales.

Ultrasound-guided axial facet joint interventions for chronic spinal pain: A narrative review.

Background: Axial facet joint interventions (e.g., medial branch block and radiofrequency ablation, facet joint intra-articular injection) are commonly performed for managing chronic spinal pain. Although traditionally performed with fluoroscopy or computed tomography (CT) guidance, ultrasound-guided techniques have also been developed for these interventions. Aims: The aim of this study is to present contemporary ultrasound-guided techniques for facet joint interventions and synthesize data addressing their accuracy, safety, and efficacy. Methods: The PubMed, MEDLINE, CINAHL, Embase, and Cochrane Central Register of Controlled Trials databases were systematically searched for studies of ultrasound-guided facet joint interventions with human subjects from November 1, 1992, to November 1, 2022. Additional sources were drawn from reference lists and citations of relevant studies. Results: We found 48 studies assessing ultrasound-guided facet joint interventions. Ultrasound guidance for injection of the cervical facet joints and their innervating nerves had favorable accuracy (78%-100%), with lower procedural time compared to fluoroscopy or CT guidance and comparable pain relief. Accuracy with ultrasound-guided lumbar facet joint intra-articular injection (86%-100%) was more reliable than medial branch block (72%-97%); analgesia was comparable to fluoroscopy and CT guidance. In general, these procedures were more challenging for patients with obesity, and deeper structures were more difficult to accurately target (e.g., lower cervical levels, L5 dorsal ramus). Conclusions: Ultrasound-guided facet joint interventions continue to evolve. Some technically challenging interventions may be impractical for widespread usage or require further technical refinement. The utility of ultrasound guidance with obesity and abnormal anatomy may be reduced.

Contexte: Les interventions sur l'articulation facettaire axiale (par exemple, le bloc de la branche médiane, l'ablation par radiofréquence et l'infiltration intra-articulaire de l'articulation facettaire sont couramment pratiquées pour traiter la douleur rachidienne chronique. Bien qu'elles soient traditionnellement réalisées sous guidage fluoroscopique ou tomodensitométrique, des techniques guidées par ultrasons ont également été mises au point pour ces interventions.Objectifs: L'objectif de cette étude est de présenter les techniques contemporaines guidées par ultrasons pour les interventions sur les articulations facettaires et de synthétiser les données relatives à ces techniques.Méthodes: Les bases de données MEDLINE, Embase et Cochrane Central Register of Controlled Trials ont fait l'objet d'une recherche systématique d'études portant sur des interventions sur les articulations facettaires guidées par ultrasons sur des sujets humains entre le 1er novembre 1992 et le 1er novembre 2022. D'autres sources ont été tirées de listes de référence et de citations d'études pertinentes.Résultats: Nous avons trouvé 48 études évaluant des interventions sur les articulations facettaires guidées par ultrasons. Le guidage par ultrasons pour l'injection des facettes cervicales et de leurs nerfs innervants avait une précision favorable (78 %-100 %) et une durée d'intervention inférieure à celle de la fluoroscopie ou de la tomodensitométrie, assorties d'un soulagement comparable de la douleur. La précision de l'injection intra-articulaire de la facette lombaire guidée par ultrasons (86 %-100 %) était plus fiable que celle du bloc de la branche médiane (72 %-97 %); l'analgésie était comparable au guidage fluoroscopique et tomodensitométrique. En général, ces procédures étaient plus difficiles pour les patients souffrant d'obésité, et les structures plus profondes étaient plus difficiles à cibler avec précision (par exemple, les niveaux cervicaux inférieurs, la branche dorsale L5).Conclusions: Les interventions sur la facette articulaire guidées par ultrasons continuent d'évoluer. Certaines interventions techniquement difficiles peuvent être impraticables pour une utilisation généralisée ou nécessiter d'autres techniques. L'utilité du guidage par ultrasons en cas d'obésité et d'anatomie anormale peut être moindre.

Effect of Fetal Delivery on Pao2/Fio2 Ratio During Mechanical Ventilation in Parturients With COVID-19: A Case Series.

It is controversial whether fetal delivery improves maternal oxygenation during mechanical ventilation. We evaluated maternal arterial partial pressure of oxygen (Pao2) to fractional oxygen concentration (Fio2) (P/F) ratios before and after delivery in this series of 15 parturients with coronavirus disease 2019 (COVID-19). Compared to the immediate postpartum period, P/F ratio was increased at 48 hours (212 ± 101 vs 271 ± 90; P = .006). Linear regression demonstrated improvement in P/F ratio during the study period (slope, 3.1; 95% confidence interval [CI], 0.87-5.34; P = .007), although predelivery and postdelivery periods separately did not exhibit any specific trend. Five patients required emergent bedside delivery. We discuss numerous considerations guiding delivery planning during mechanical ventilation.

Extracorporeal Membrane Oxygenation for Pregnant and Postpartum Patients.

Extracorporeal membrane oxygenation (ECMO) has seen increasing use for critically ill pregnant and postpartum patients over the past decade. Growing experience continues to demonstrate the feasibility of ECMO in obstetric patients and attest to its favorable outcomes. However, the interaction of pregnancy physiology with ECMO life support requires careful planning and adaptation for success. Additionally, the maintenance of fetal oxygenation and perfusion is essential for safely continuing pregnancy during ECMO support. This review summarizes the considerations for use of ECMO in obstetric patients and how to address these concerns.

Presentation and aetiology of paediatric trigger finger: a systematic review.

Paediatric trigger finger is a rare condition distinct from paediatric trigger thumb and adult trigger digits. We performed a systematic review of paediatric trigger finger presentation and aetiology in order to guide workup and management. Fifty-one studies with 193 patients and 398 trigger fingers were included. Most patients had a single, unilateral trigger finger (54%). Fifty-five patients (29%) had an underlying condition, such as mucopolysaccharidosis; these cases appeared to be associated with multiple or bilateral trigger fingers or with carpal tunnel syndrome. All patients with mucopolysaccharidosis were treated surgically. Conservative management was reported in 33% of all patients, and two-thirds of these did not need further intervention. Patients undergoing surgical release infrequently had recurrence of triggering (6%). We propose an algorithmic approach for patients presenting with paediatric trigger finger. Presence of bilateral or multiple trigger digits or concomitant carpal tunnel syndrome should raise suspicion for an atypical underlying pathology.

Not All Subarachnoid Hemorrhages in Pregnancy Are Intracranial in Origin: A Case Report.

Subarachnoid hemorrhage is uncommon in pregnancy and may be caused by intracranial aneurysms, arteriovenous malformations, venous thrombosis, or preeclampsia. We present an unusual case of subarachnoid hemorrhage in a term parturient where the bleeding originated from an extracranial source, namely a cervicothoracic arteriovenous malformation. This case highlights the challenge of diagnosing this condition when the initial presentation may be nonspecific, lacking in neurologic deficits, and confounded by the simultaneous presence of preeclampsia.

Hsp70 and DNAJA2 limit CFTR levels through degradation.

Cystic Fibrosis is caused by mutations in the CFTR anion channel, many of which cause its misfolding and degradation. CFTR folding depends on the Hsc70 and Hsp70 chaperones and their co-chaperone DNAJA1, but Hsc70/Hsp70 is also involved in CFTR degradation. Here, we address how these opposing functions are balanced. DNAJA2 and DNAJA1 were both important for CFTR folding, however overexpressing DNAJA2 but not DNAJA1 enhanced CFTR degradation at the endoplasmic reticulum by Hsc70/Hsp70 and the E3 ubiquitin ligase CHIP. Excess Hsp70 also promoted CFTR degradation, but this occurred through the lysosomal pathway and required CHIP but not complex formation with HOP and Hsp90. Notably, the Hsp70 inhibitor MKT077 enhanced levels of mature CFTR and the most common disease variant ΔF508-CFTR, by slowing turnover and allowing delayed maturation, respectively. MKT077 also boosted the channel activity of ΔF508-CFTR when combined with the corrector compound VX809. Thus, the Hsp70 system is the major determinant of CFTR degradation, and its modulation can partially relieve the misfolding phenotype.

Supplemental perioperative intravenous crystalloids for postoperative nausea and vomiting.

BACKGROUND: Postoperative nausea and vomiting (PONV) is a common complication following general anaesthesia. It may be associated with patient dissatisfaction, increased costs of treatment, and unintended admission to hospital.Supplemental intravenous crystalloid administration in the perioperative period may be a simple intervention to prevent PONV. OBJECTIVES: To assess whether supplemental intravenous crystalloid administration prevents PONV in patients undergoing surgical procedures under general anaesthesia. SEARCH METHODS: We searched the Cochrane Central Register of Controlled Trials (CENTRAL; 2018, Issue 7), MEDLINE (1946 to August 2018), Embase (1947 to August 2018), and the Cumulative Index of Nursing and Allied Health Literature (CINAHL; 1971 to August 2018). We searched clinical trials registers for ongoing or unpublished completed studies (August 2018), handsearched three major journals (British Journal of Anaesthesia, European Journal of Anaesthesiology, and Anesthesiology; August 2018), and conducted backward and forward citation searching of relevant articles. SELECTION CRITERIA: We included randomized controlled trials of participants older than six months undergoing surgical procedures under general anaesthesia and given supplemental perioperative intravenous crystalloids, defined as a volume larger than that received by a comparator group, to prevent PONV. DATA COLLECTION AND ANALYSIS: We used the standard methodological procedures described by Cochrane. MAIN RESULTS: We included 41 studies (4224 participants). Participants underwent ambulatory or short length of stay surgical procedures, and were predominantly American Society of Anesthesiology (ASA) class I or II. There is one study awaiting classification and three ongoing studies. All studies took place in surgical centres, and were conducted in geographically diverse settings. Risk of bias was generally unclear across all domains.Supplemental intravenous crystalloid administration probably reduces the cumulative risk of postoperative nausea (PON) (risk ratio (RR) 0.62, 95% confidence interval (CI) 0.51 to 0.75; 18 studies; 1766 participants; moderate-certainty evidence). When the postoperative period was divided into early (first six hours postoperatively) and late (at the time point closest to or including 24 hours postoperatively) time points, the intervention reduced the risk of early PON (RR 0.67, 95% CI 0.58 to 0.78; 20 studies; 2310 participants; moderate-certainty evidence) and late PON (RR 0.47, 95% CI 0.32 to 0.69; 17 studies; 1682 participants; moderate-certainty evidence).Supplemental intravenous crystalloid administration probably reduces the risk of postoperative vomiting (POV) (RR 0.50, 95% CI 0.40 to 0.63; 20 studies; 1970 participants; moderate-certainty evidence). The intervention specifically reduced both early POV (RR 0.56, 95% CI 0.41 to 0.76; 19 studies; 1998 participants; moderate-certainty evidence) and late POV (RR 0.48, 95% CI 0.29 to 0.79; 15 studies; 1403 participants; moderate-certainty evidence).Supplemental intravenous crystalloid administration probably reduces the need for pharmacologic treatment of PONV (RR 0.62, 95% CI 0.51 to 0.76; 23 studies; 2416 participants; moderate-certainty evidence).The effect of supplemental intravenous crystalloid administration on the risk of unplanned postoperative admission to hospital is unclear (RR 1.05, 95% CI 0.77 to 1.43; 3 studies; 235 participants; low-certainty evidence).No studies reported serious adverse events that may occur following supplemental perioperative intravenous crystalloid administration (i.e. admission to high-dependency unit, postoperative cardiac or respiratory complication, or death). AUTHORS' CONCLUSIONS: There is moderate-certainty evidence that supplemental perioperative intravenous crystalloid administration reduces PON and POV, in ASA class I to II patients receiving general anaesthesia for ambulatory or short length of stay surgical procedures. The intervention probably also reduces the risk of pharmacologic treatment for PONV. The effect of the intervention on the risk of unintended postoperative admission to hospital is unclear. The risk of serious adverse events resulting from supplemental perioperative intravenous crystalloid administration is unknown as no studies reported this outcome. The one study awaiting classification may alter the conclusions of the review once assessed.

Biphasic concentration-dependent interaction between imidacloprid and dietary phytochemicals in honey bees (Apis mellifera).

BACKGROUND: The presence of the neonicotinoid imidacloprid in nectar, honey, pollen, beebread and beeswax has been implicated in declines worldwide in the health of the western honey bee Apis mellifera. Certain phytochemicals, including quercetin and p-coumaric acid, are ubiquitous in the honey bee diet and are known to upregulate cytochrome P450 genes encoding enzymes that detoxify insecticides. Thus, the possibility exists that these dietary phytochemicals interact with ingested imidacloprid to ameliorate toxicity by enhancing its detoxification. APPROACH: Quercetin and p-coumaric acid were incorporated in a phytochemical-free artificial diet individually and together along with imidacloprid at a range of field-realistic concentrations. In acute toxicity bioassays, honey bee 24- and 48- hour imidacloprid LC50 values were determined in the presence of the phytochemicals. Additionally, chronic toxicity bioassays were conducted using varying concentrations of imidacloprid in diets with the phytochemicals to test impacts of phytochemicals on longevity. RESULTS: In acute toxicity bioassays, the phytochemicals had no effect on imidacloprid LC50 values. In chronic toxicity longevity bioassays, phytochemicals enhanced honey bee survival at low imidacloprid concentrations (15 and 45 ppb) but had a negative effect at higher concentrations (105 ppb and 135 ppb). p-Coumaric acid alone increased honey bee longevity at concentrations of 15 and 45 ppb imidacloprid (hazard ratio (HR): 0.83 and 0.70, respectively). Quercetin alone and in combination with p-coumaric acid similarly enhanced longevity at 45 ppb imidacloprid (HR:0.81 and HR:0.77, respectively). However, p-coumaric acid in combination with 105 ppb imidacloprid and quercetin in combination with 135 ppb imidacloprid increased honey bee HR by approximately 30% (HR:1.33 and HR:1.30, respectively). CONCLUSIONS: The biphasic concentration-dependent response of honey bees to imidacloprid in the presence of two ubiquitous dietary phytochemicals indicates that there are limits to the protective effects of the natural diet of honey bees against neonicotinoids based on their own inherent toxicity.

Structures of ubiquitin-like (Ubl) and Hsp90-like domains of sacsin provide insight into pathological mutations.

Autosomal recessive spastic ataxia of Charlevoix-Saguenay (ARSACS) is a neurodegenerative disease that is caused by mutations in the SACS gene. The product of this gene is a very large 520-kDa cytoplasmic protein, sacsin, with a ubiquitin-like (Ubl) domain at the N terminus followed by three large sacsin internal repeat (SIRPT) supradomains and C-terminal J and HEPN domains. The SIRPTs are predicted to contain Hsp90-like domains, suggesting a potential chaperone activity. In this work, we report the structures of the Hsp90-like Sr1 domain of SIRPT1 and the N-terminal Ubl domain determined at 1.55- and 2.1-Å resolutions, respectively. The Ubl domain crystallized as a swapped dimer that could be relevant in the context of full-length protein. The Sr1 domain displays the Bergerat protein fold with a characteristic nucleotide-binding pocket, although it binds nucleotides with very low affinity. The Sr1 structure reveals that ARSACS-causing missense mutations (R272H, R272C, and T201K) disrupt protein folding, most likely leading to sacsin degradation. This work lends structural support to the view of sacsin as a molecular chaperone and provides a framework for future studies of this protein.

The Effect of Written Information on Recall of Surgical Risks of Carpal Tunnel Release Surgery: A Randomized Controlled Study.

BACKGROUND: Written information has been thought to help patients recall surgical risks discussed during the informed consent process, but has not been assessed for carpal tunnel release, a procedure with the rare but serious risk of complex regional pain syndrome. The authors' objective was to determine whether providing a pamphlet would improve patients' ability to remember the risks of surgery. METHODS: Sixty patients seen for carpal tunnel release were included in this prospective, single-blind, randomized study. Patients received either a written pamphlet of the risks of surgery or no additional information following a standardized consultation. Two weeks after the initial consultation, patients were contacted to assess their risk recall and whether they had read about the operation from any source. RESULTS: There was no difference in terms of the number of risks recalled between pamphlet (1.33 ± 1.21) or control groups (1.45 ± 1.22; p = 0.73). Recall of infection was better in the pamphlet group (p < 0.05). No patients remembered complex regional pain syndrome. There was no difference in the proportion of people who read additional information about carpal tunnel release surgery between the pamphlet (34.8 percent) and control groups (21.4 percent; p = 0.39), but reading about carpal tunnel release surgery was associated with improved recall (2.45 ± 1.13 versus 0.77 ± 0.91; p < 0.01). CONCLUSIONS: Reading about surgery improved risk recall, but providing this information in the form of a pamphlet did not, nor did it affect patients' ability to recall the risk of complex regional pain syndrome. These results demonstrate that surgeons should implement additional measures to improve comprehension of surgical risks. CLINICAL QUESTION/LEVEL OF EVIDENCE: Therapeutic, I.

Shared medical appointments as a new model for carpal tunnel surgery consultation: A randomized clinical trial.

BACKGROUND: In chronic disease management, shared medical appointments have been shown to improve clinic access, productivity and patient education. However, adoption of this model in surgical consultation is limited, and its effect on surgical patients' satisfaction, comfort and surgical risk recall is unknown. OBJECTIVE: To determine whether shared medical appointments could be applied to carpal tunnel surgery consultation while being equally effective as individual consultation for risk recall, patient comfort and satisfaction. METHODS: A prospective randomized trial involving 80 patients referred for carpal tunnel release consultation, in which patients were assigned to an educational discussion individually or as part of a shared appointment, was conducted. In a blinded fashion, patients were contacted preoperatively to assess their risk recall and postoperatively to rate their overall satisfaction, comfort and satisfaction with the surgeon. RESULTS: Patient demographics were equal. Surgical risk recall was equivalent between shared and individual consults (2.06±1.15 versus 1.64±1.04; P=0.11). More participants in the shared appointments condition remembered the specific risks of infection (61.1% versus 33.3%; P=0.020) and bleeding (30.6% versus 10.3%; P=0.028). There was no difference in overall satisfaction (8.70 versus 8.88; P=0.75), satisfaction with the surgeon (8.05 versus 8.13; P=0.92) or overall comfort (8.80 versus 8.31; P=0.46). DISCUSSION: Shared medical appointments for carpal tunnel surgery consultation were equivalent to individual consultation in terms of surgical risk recall, patient satisfaction and comfort. CONCLUSION: These results support the use of shared appointments for large-volume, low-variation surgery.

HISTORIQUE: Dans la prise en charge des maladies chroniques, les rendez-vous médicaux groupés améliorent l'accès aux cliniques, la productivité et l'éducation des patients. Cependant, ce modèle est peu utilisé en consultation chirurgicale, et on n'en connaît pas l'effet sur la satisfaction des patients en voie d'être opérés, leur confort et leurs souvenirs des risques de la chirurgie. OBJECTIF: Déterminer si les rendez-vous médicaux groupés peuvent être utilisés dans le cadre des consultations avant une chirurgie du canal carpien avec la même efficacité qu'une consultation individuelle sur le plan des souvenirs des risques, du confort et de la satisfaction des patients. MÉTHODOLOGIE: Les chercheurs ont réalisé un essai aléatoire et prospectif auprès de 80 patients dirigés vers une consultation en vue d'une libération du canal carpien. Les patients ont été affectés à une séance d'éducation individuelle ou à un rendez-vous groupé. À l'aveugle, on a communiqué avec les patients avant l'opération pour évaluer leurs souvenirs des risques et après l'opération pour déterminer leur satisfaction et leur confort globaux ainsi que leur satisfaction envers le chirurgien. RÉSULTATS: Les patients avaient une démographie similaire. Leurs souvenirs des risques de la chirurgie étaient similaires dans les consultations groupées ou individuelles (2,06±1,15 par rapport à 1,64±1,04; P=0,11). Plus de patients des consultations groupées se souvenaient des risques spécifiques d'infection (61,1 % par rapport à 33,3 %; P=0,020) et de saignements (30,6 % par rapport à 10,3 %; P=0,028). Il n'y avait pas de différence dans la satisfaction globale (8,70 par rapport à 8,88; P=0,75), la satisfaction envers le chirurgien (8,05 par rapport à 8,13; P=0,92) ou le confort global (8,80 par rapport à 8,31; P=0,46). EXPOSÉ: Les rendez-vous médicaux groupés lors des consultations en vue d'une chirurgie du canal carpien étaient équivalents aux consultations individuelles pour ce qui est du souvenir des risques chirurgicaux, de la satisfaction des patients et du confort. CONCLUSION: Ces résultats appuient l'utilisation des rendez-vous groupés pour les opérations peu variables et à grand volume.

Heat Shock Protein 90 kDa (Hsp90) Has a Second Functional Interaction Site with the Mitochondrial Import Receptor Tom70.

To accomplish its crucial role, mitochondria require proteins that are produced in the cytosol, delivered by cytosolic Hsp90, and translocated to its interior by the translocase outer membrane (TOM) complex. Hsp90 is a dimeric molecular chaperone and its function is modulated by its interaction with a large variety of co-chaperones expressed within the cell. An important family of co-chaperones is characterized by the presence of one TPR (tetratricopeptide repeat) domain, which binds to the C-terminal MEEVD motif of Hsp90. These include Tom70, an important component of the TOM complex. Despite a wealth of studies conducted on the relevance of Tom70·Hsp90 complex formation, there is a dearth of information regarding the exact molecular mode of interaction. To help fill this void, we have employed a combined experimental strategy consisting of cross-linking/mass spectrometry to investigate binding of the C-terminal Hsp90 domain to the cytosolic domain of Tom70. This approach has identified a novel region of contact between C-Hsp90 and Tom70, a finding that is confirmed by probing the corresponding peptides derived from cross-linking experiments via isothermal titration calorimetry and mitochondrial import assays. The data generated in this study are combined to input constraints for a molecular model of the Hsp90/Tom70 interaction, which has been validated by small angle x-ray scattering, hydrogen/deuterium exchange, and mass spectrometry. The resultant model suggests that only one of the MEEVD motifs within dimeric Hsp90 contacts Tom70. Collectively, our findings provide significant insight on the mechanisms by which preproteins interact with Hsp90 and are translocated via Tom70 to the mitochondria.

Hsp70 cochaperones HspBP1 and BAG-1M differentially regulate steroid hormone receptor function.

Hsp70 binding protein 1 (HspBP1) and Bcl2-associated athanogene 1 (BAG-1), the functional orthologous nucleotide exchange factors of the heat shock protein 70 kilodalton (Hsc70/Hsp70) chaperones, catalyze the release of ADP from Hsp70 while inducing different conformational changes of the ATPase domain of Hsp70. An appropriate exchange rate of ADP/ATP is crucial for chaperone-dependent protein folding processes. Among Hsp70 client proteins are steroid receptors such as the glucocorticoid receptor (GR), the mineralocorticoid receptor (MR), and the androgen receptor (AR). BAG-1 diversely affects steroid receptor activity, while to date the influence of HspBP1 on steroid receptor function is mostly unknown. Here, we compared the influence of HspBP1 and BAG-1M on Hsp70-mediated steroid receptor folding complexes and steroid receptor activity. Coimmunoprecipitation studies indicated preferential binding of Hsp40 and the steroid receptors to BAG-1M as compared to HspBP1. Furthermore, Hsp70 binding to the ligand-binding domain of GR was reduced in the presence of HspBP1 but not in the presence of BAG-1M as shown by pull-down assays. Reporter gene experiments revealed an inhibitory effect on GR, MR, and AR at a wide range of HspBP1 protein levels and at hormone concentrations at or approaching saturation. BAG-1M exhibited a transition from stimulatory effects at low BAG-1M levels to inhibitory effects at higher BAG-1M levels. Overall, BAG-1M and HspBP1 had differential impacts on the dynamic composition of steroid receptor folding complexes and on receptor function with important implications for steroid receptor physiology.

Identification of an allosteric pocket on human hsp70 reveals a mode of inhibition of this therapeutically important protein.

Hsp70s are important cancer chaperones that act upstream of Hsp90 and exhibit independent anti-apoptotic activities. To develop chemical tools for the study of human Hsp70, we developed a homology model that unveils a previously unknown allosteric site located in the nucleotide binding domain of Hsp70. Combining structure-based design and phenotypic testing, we discovered a previously unknown inhibitor of this site, YK5. In cancer cells, this compound is a potent and selective binder of the cytosolic but not the organellar human Hsp70s and has biological activity partly by interfering with the formation of active oncogenic Hsp70/Hsp90/client protein complexes. YK5 is a small molecule inhibitor rationally designed to interact with an allosteric pocket of Hsp70 and represents a previously unknown chemical tool to investigate cellular mechanisms associated with Hsp70.

The DNAJA2 substrate release mechanism is essential for chaperone-mediated folding.

DNAJA1 (DJA1/Hdj2) and DNAJA2 (DJA2) are the major J domain partners of human Hsp70/Hsc70 chaperones. Although they have overall similarity with the well characterized type I co-chaperones from yeast and bacteria, they are biologically distinct, and their functional mechanisms are poorly characterized. We identified DJA2-specific activities in luciferase folding and repression of human ether-a-go-go-related gene (HERG) trafficking that depended on its expression levels in cells. Mutations in different internal domains of DJA2 abolished these effects. Using purified proteins, we addressed the mechanistic defects. A mutant lacking the region between the zinc finger motifs (DJA2-Δm2) was able to bind substrate similar to wild type but was incapable of releasing substrate during its transfer to Hsc70. The equivalent mutation in DJA1 also abolished its substrate release. A DJA2 mutant (DJA-221), which had its C-terminal dimerization region replaced by that of DJA1, was inactive but retained its ability to release substrate. The release mechanism required the J domain and ATP hydrolysis by Hsc70, although the nucleotide dependence diverged between DJA2 and DJA1. Limited proteolysis suggested further conformational differences between the two wild-type co-chaperones and the mutants. Our results demonstrate an essential role of specific DJA domains in the folding mechanism of Hsc70.

Interaction between the human mitochondrial import receptors Tom20 and Tom70 in vitro suggests a chaperone displacement mechanism.

The mitochondrial import receptor Tom70 contains a tetratricopeptide repeat (TPR) clamp domain, which allows the receptor to interact with the molecular chaperones, Hsc70/Hsp70 and Hsp90. Preprotein recognition by Tom70, a critical step to initiate import, is dependent on these cytosolic chaperones. Preproteins are subsequently released from the receptor for translocation across the outer membrane, yet the mechanism of this step is unknown. Here, we report that Tom20 interacts with the TPR clamp domain of Tom70 via a conserved C-terminal DDVE motif. This interaction was observed by cross-linking endogenous proteins on the outer membrane of mitochondria from HeLa cells and in co-precipitation and NMR titrations with purified proteins. Upon mutation of the TPR clamp domain or deletion of the DDVE motif, the interaction was impaired. In co-precipitation experiments, the Tom20-Tom70 interaction was inhibited by C-terminal peptides from Tom20, as well as from Hsc70 and Hsp90. The Hsp90-Tom70 interaction was measured with surface plasmon resonance, and the same peptides inhibited the interaction. Thus, Tom20 competes with the chaperones for Tom70 binding. Interestingly, antibody blocking of Tom20 did not increase the efficiency of Tom70-dependent preprotein import; instead, it impaired the Tom70 import pathway in addition to the Tom20 pathway. The functional interaction between Tom20 and Tom70 may be required at a later step of the Tom70-mediated import, after chaperone docking. We suggest a novel model in which Tom20 binds Tom70 to facilitate preprotein release from the chaperones by competition.

Hsp40 chaperones promote degradation of the HERG potassium channel.

Loss of function mutations in the hERG (human ether-a-go-go related gene or KCNH2) potassium channel underlie the proarrhythmic cardiac long QT syndrome type 2. Most often this is a consequence of defective trafficking of hERG mutants to the cell surface, with channel retention and degradation at the endoplasmic reticulum. Here, we identify the Hsp40 type 1 chaperones DJA1 (DNAJA1/Hdj2) and DJA2 (DNAJA2) as key modulators of hERG degradation. Overexpression of the DJAs reduces hERG trafficking efficiency, an effect eliminated by the proteasomal inhibitor lactacystin or with DJA mutants lacking their J domains essential for Hsc70/Hsp70 activation. Both DJA1 and DJA2 cause a decrease in the amount of hERG complexed with Hsc70, indicating a preferential degradation of the complex. Similar effects were observed with the E3 ubiquitin ligase CHIP. Both the DJAs and CHIP reduce hERG stability and act differentially on folding intermediates of hERG and the disease-related trafficking mutant G601S. We propose a novel role for the DJA proteins in regulating degradation and suggest that they act at a critical point in secretory pathway quality control.