Microbial transformation of dietary xenobiotics shapes gut microbiome composition.

Diet is a major determinant of gut microbiome composition, and variation in diet-microbiome interactions may contribute to variation in their health consequences. To mechanistically understand these relationships, here we map interactions between ∼150 small-molecule dietary xenobiotics and the gut microbiome, including the impacts of these compounds on community composition, the metabolic activities of human gut microbes on dietary xenobiotics, and interindividual variation in these traits. Microbial metabolism can toxify and detoxify these compounds, producing emergent interactions that explain community-specific remodeling by dietary xenobiotics. We identify the gene and enzyme responsible for detoxification of one such dietary xenobiotic, resveratrol, and demonstrate that this enzyme contributes to interindividual variation in community remodeling by resveratrol. Together, these results systematically map interactions between dietary xenobiotics and the gut microbiome and connect toxification and detoxification to interpersonal differences in microbiome response to diet.

A4 Thoracolumbar Fracture Class Is Associated With a Greater Degree of Vertebral Height Loss in Conservatively Managed Patients.

INTRODUCTION: Thoracolumbar (TL) junction fractures are common, often resulting from high-energy trauma or osteoporosis, and may lead to neurological deficits, deformities, or chronic pain. Treatment decisions for neurologically intact patients remain controversial, with nonsurgical management often favored. The AO classification system has been used to characterize thoracolumbar fractures using fracture morphology and clinical factors affecting clinical decision-making for fracture management. This study aims to assess the radiographic outcomes of utilizing a thoracolumbosacral orthosis (TLSO) brace in neurologically intact patients with TL fractures based on the AO classification system. METHODS: A retrospective analysis of 43 patients was conducted using data from the VCU Spine Database on patients with TL fractures managed conservatively with a TLSO brace from 2010 to 2019. Demographic variables and radiographic measurements of anterior height loss were analyzed and stratified by AO fracture class. RESULTS: Significant differences were observed in anterior height loss between AO fracture classes, with A4 fractures showing significantly greater anterior height loss at initial presentation (27.6 + 4.8%) compared to A1/A2 (16.1 + 2.2%; p=0.049). At follow up, A4 fractures had a significantly greater anterior height loss (40.2 + 6.6%) than both the A1/A2 (22.4 + 2.9%; p=0.029) and A3 fracture classes (20.5 + 3.6; p=0.020). CONCLUSIONS: The study highlights significant differences in anterior height loss among AO fracture classes, suggesting varying degrees of severity and potential implications for clinical management. While conservative treatment with TLSO braces may provide pain relief, surgical intervention may offer better structural recovery, especially in more severe fractures. Conservative management of TL fractures with TLSO braces may result in greater anterior height loss, particularly in A4 fractures, emphasizing the need for individualized treatment decisions. Further research, including prospective studies, is warranted to validate these findings and guide clinical practice effectively.

The systemic complexity of a monogenic disease: the molecular network of spinal muscular atrophy.

Monogenic diseases are well-suited paradigms for the causal analysis of disease-driving molecular patterns. Spinal Muscular Atrophy (SMA) is one such monogenic model caused by mutation or deletion of the Survival of motor neuron 1 (SMN1) gene. Although several functions of the SMN protein have been studied, single functions and pathways alone do not allow to identify critical disease-driving molecules. Here, we analyzed the systemic characteristics of SMA employing proteomics, phosphoproteomics, translatomics and interactomics from two mouse models with different disease-severities and genetics. This systems approach revealed sub-networks and proteins characterizing commonalities and differences of both models. To link the identified molecular networks with the disease-causing SMN protein, we combined SMN-interactome data with both proteomes creating a comprehensive representation of SMA. By this approach, disease hubs and bottlenecks between SMN and downstream pathways could be identified. Linking a disease-causing molecule with widespread molecular dysregulations via multiomics is a concept for analyses of monogenic diseases.

What could be the function of the spinal muscular atrophy-causing protein SMN in macrophages?

Spinal Muscular Atrophy (SMA), a neurodegenerative disorder, extends its impact beyond the nervous system. The central protein implicated in SMA, Survival Motor Neuron (SMN) protein, is ubiquitously expressed and functions in fundamental processes such as alternative splicing, translation, cytoskeletal dynamics and signaling. These processes are relevant for all cellular systems, including cells of the immune system such as macrophages. Macrophages are capable of modulating their splicing, cytoskeleton and expression profile in order to fulfil their role in tissue homeostasis and defense. However, less is known about impairment or dysfunction of macrophages lacking SMN and the subsequent impact on the immune system of SMA patients. We aimed to review the potential overlaps between SMN functions and macrophage mechanisms highlighting the need for future research, as well as the current state of research addressing the role of macrophages in SMA.

From data to discovery: AI-guided analysis of disease-relevant molecules in spinal muscular atrophy (SMA).

Spinal Muscular Atrophy is caused by partial loss of survival of motoneuron (SMN) protein expression. The numerous interaction partners and mechanisms influenced by SMN loss result in a complex disease. Current treatments restore SMN protein levels to a certain extent, but do not cure all symptoms. The prolonged survival of patients creates an increasing need for a better understanding of SMA. Although many SMN-protein interactions, dysregulated pathways, and organ phenotypes are known, the connections among them remain largely unexplored. Monogenic diseases are ideal examples for the exploration of cause-and-effect relationships to create a network describing the disease-context. Machine learning tools can utilize such knowledge to analyze similarities between disease-relevant molecules and molecules not described in the disease so far. We used an artificial intelligence-based algorithm to predict new genes of interest. The transcriptional regulation of 8 out of 13 molecules selected from the predicted set were successfully validated in an SMA mouse model. This bioinformatic approach, using the given experimental knowledge for relevance predictions, enhances efficient targeted research in SMA and potentially in other disease settings.

Validation of the RSClin risk calculator in the National Cancer Data Base.

BACKGROUND: Guidelines recommend the use of genomic assays such as OncotypeDx to aid in decisions regarding the use of chemotherapy for hormone receptor-positive, HER2-negative (HR+/HER2-) breast cancer. The RSClin prognostic tool integrates OncotypeDx and clinicopathologic features to predict distant recurrence and chemotherapy benefit, but further validation is needed before broad clinical adoption. METHODS: This study included patients from the National Cancer Data Base (NCDB) who were diagnosed with stage I-III HR+/HER2- breast cancer from 2010 to 2020 and received adjuvant endocrine therapy with or without chemotherapy. RSClin-predicted chemotherapy benefit was stratified into low (<3% reduction in distant recurrence), intermediate (3%-5%), and high (>5%). Cox models were used to model mortality adjusted for age, comorbidity index, insurance, and race/ethnicity. RESULTS: A total of 285,441 patients were identified for inclusion from the NCDB, with an average age of 60 years and a median follow-up of 58 months. Chemotherapy was associated with improved overall survival only for those predicted to have intermediate (adjusted hazard ratio [aHR], 0.68; 95% confidence interval [CI], 0.60-0.79) and high benefit per RSClin (aHR, 0.66; 95% CI, 0.61-0.72). Consistent benefit was seen in the subset with a low OncotypeDx score (<26) and intermediate (aHR, 0.66; 95% CI, 0.53-0.82) or high (aHR, 0.71; 95% CI, 0.58-0.86) RSClin-predicted benefit. No survival benefit with chemotherapy was seen in patients with a high OncotypeDx score (≥26) and low benefit per RSClin (aHR, 1.70; 95% CI, 0.41-6.99). CONCLUSIONS: RSClin may identify high-risk patients who benefit from treatment intensification more accurately than OncotypeDx, and further prospective study is needed.

The effect of poor glycemic control on cognitive function in children and adolescents with type 1 diabetes mellitus: A single-center cross-sectional study (2019-2020).

OBJECTIVES: To investigate the effect of chronic hyperglycemia, hypoglycemia, and diabetic ketoacidosis (DKA) on the cognitive function of children and adolescents with type 1 diabetes mellitus (T1DM), and explore whether early disease onset correlated with cognitive impairment. METHODS: Children and adolescents with T1DM who attended the Pediatric Diabetes Clinic between January 2019 and 2020, aged between 5-14 years, with a mean of 3 glycated hemoglobin (HbA1c) readings ≥7% (53 mmol/mol) during the study period reflecting a hyperglycemic status and who agreed to participate were interviewed about their history of hypoglycemia and DKA. Participants were personally interviewed by a clinical psychologist to assess their cognitive function using a validated Arabic version of the Stanford-Binet test. RESULTS: Higher mean HbA1c levels were associated with cognitive dysfunction in three verbal domains: fluid reasoning, quantitative reasoning, and working memory. Frequent hypoglycemia negatively affected verbal knowledge. In contrast, significant hypoglycemia affected both verbal and nonverbal domains of cognition, specifically verbal and nonverbal fluid reasoning, knowledge, and working memory. Children with recurrent DKA performed below average in nonverbal fluid reasoning tasks. Additionally, moderate or severe DKA, regardless of its frequency, affected children's overall intelligence quotient. CONCLUSION: Regardless of disease onset, exposure to glycemic variability subjects children and adolescents to subtle and measurable cognitive dysfunction resulting in significant morbidity.

A cooperation experiment with white-handed gibbons (Hylobates lar).

Cooperative behaviors among individuals of numerous species play a crucial role in social interactions. There is a special interest in investigating the occurrence of cooperation among apes because this knowledge could also shed light on evolutionary processes and help us understand the origin and development of cooperation in humans and primates in general. Gibbons are phylogenetically intermediate between the great apes and monkeys, and therefore represent a unique opportunity for comparisons. The aim of the present study was to discover whether or not white-handed gibbons (Hylobates lar) show cooperative behaviors. In order to test for the respective behaviors, the gibbons were presented with a commonly used experimental cooperative rope-pulling task. The gibbons in this study did not exhibit cooperative behaviors during the problem-solving task. However, prior training procedures could not be fully completed, hence this project constitutes only the onset of exploring cooperative behaviors in gibbons. Additional behavioral observations revealed that the gibbons spent significantly more time "out of arm's reach to everyone", suggesting that they are less often involved in social interactions, than other, more cooperative primates.

Regarding the Influence of Additives and Additional Plasma-Induced Chemical Ionization on Adduct Formation in ESI/IMS/MS.

Ion mobility spectrometers (IMS) separate ions based on their ion mobility, which depends mainly on collision cross-section, mass, and charge of the ions. However, the performance is often hampered in electrospray ionization (ESI) by the appearance of multiple ion mobility peaks in the spectrum for the same analyte due to clustering and additional sodium adducts. In this work, we investigate the influence of solvents and buffer additives on the detected ion mobility peaks using ESI. Additionally, we investigate the effects of an additional chemical ionization (CI) induced by plasma ionization on the ions formed by electrospray. For this purpose, we coupled our high-resolution IMS with a resolving power of Rp = 100 to a time-of-flight mass spectrometer. Depending on the analyte and the chosen additives, the ionization process can be influenced during the electrospray process. For the herbicide isoproturon, the addition of 5 mM sodium acetate results in the formation of the sodium adduct [M + Na]+, which is reflected in the ion mobility K0 of 1.22 cm2/(V·s). In contrast, the addition of 5 mM ammonium acetate yields the protonated species [M + H]+ and a correspondingly higher K0 of 1.29 cm2/(V·s). In some cases, as with the herbicide pyrimethanil, the addition of sodium acetate can completely suppress ionizations. By carefully choosing the solvent additive for ESI-IMS or additional CI, the formation of different ion mobility peaks can be observed. This can facilitate the assignment of ions to ion mobility peaks using IMS as a compact, stand-alone instrument, e.g., for on-site analysis.

Outpatient Parenteral Antibiotic Therapy in Older Adults.

Outpatient parenteral antimicrobial therapy (OPAT) for older adults is a complex process that involves multiple stakeholders and care coordination, but it is a useful and patient-centered tool with opportunities for the treatment of complicated infections, improved patient satisfaction, and reduced health-care costs. Older age should not be an exclusion for OPAT but rather prompt the OPAT provider to thoroughly evaluate candidacy and safety. Amid the on-going COVID-19 pandemic, innovations in OPAT are needed to shepherd OPAT care into a more patient-centered, thoughtful practice, whereas minimizing harm to older patients from unnecessary health-care exposure and thus health-care associated infections.

A Promising Diagnostic Role of Immunohistochemical Expression of Insulin-Like Growth Factor II mRNA Binding Protein 3 (IMP3) in Pancreatic Lesions Using Endoscopic Ultrasound-Guided Fine Needle Aspiration (EUS-FNA) Cytology.

BACKGROUND: Poor prognosis and short survival of patients harboring pancreatic cancer emerge how advanced disease it is. In a trial to achieve the earliest and most accurate diagnosis to manage this progressive disease, we proposed that using endoscopic ultrasound-guided fine needle aspiration (EUS-FNA) with an adjuvant diagnostic immunohistochemical marker would give better diagnostic results. IMP3 has gained recently wide attention, as many studies found that IMP3 has not only diagnostic but also prognostic role in different types of malignancies. AIM OF THE STUDY: This prospective work is to assess the diagnostic role of EUS-FNA combined with the immunohistochemical expression of IMP3 on different benign and malignant pancreatic lesions. MATERIAL AND METHOD: The included pancreatic lesions (n = 140) were obtained by EUS-FNA technique and stained for IMP3 immunohistochemically. Paraffin blocks from patients who underwent excision (n = 92) or core biopsies (n = 48) were performed for confirming diagnosis. RESULTS: The combined method for diagnosis showed that IMP3 was positive in 78.7%, 91.7%, 100% PAC, mucinous neoplasm with high grade dysplasia, and IPMN with high grade dysplasia, respectively, while almost all benign lesions showed negative IMP3. Also, this method showed sensitivity (78.26%), specificity (95.83%), and accuracy (84.3%). CONCLUSION: EUS-FNA cytology with IMP3 could be a reliable diagnostic tool especially for assessment of malignant pancreatic lesions.

The association between comorbidities and coronavirus disease 2019 hospitalization among people with HIV differs by age.

OBJECTIVES: To determine whether factors associated with coronavirus disease 2019 (COVID-19) hospitalization among people with HIV (PWH) differ by age stratum. DESIGN: Retrospective cohort study. METHODS: All adult PWH with a positive SARS-CoV-2 PCR in a public safety-net health system between 1 March 2020 and 28 February 2021 and a Veterans Affairs Medical Center between 1 1 March 2020 and 15 November 2020 in Atlanta, Georgia were included. We performed multivariable logistic regression to determine demographic and clinical factors associated with COVID-19 hospitalization overall and stratified by age less than 50 and at least 50 years. RESULTS: Three hundred and sixty-five PWH (mean age 49 years, 74% cisgender male, 82% black) were included. Ninety-six percent were on antiretroviral therapy (ART), 87% had CD4 + T-cell count at least 200 cells/µl, and 89% had HIV-1 RNA less than 200 copies/ml. Overall, age [adjusted odds ratio (aOR) 95% confidence interval (CI) 1.07 (1.04-1.10)], later date of SARS-CoV-2 infection [aOR 0.997 (0.995-1.00)], heart disease [aOR 2.27 (1.06-4.85)], and history of hepatitis C virus (HCV) [aOR 2.59 (1.13-5.89)] were associated with COVID-19 hospitalization. Age-adjusted comorbidity burden was associated with 30% increased risk of hospitalization [aOR 1.30 (1.11-1.54)]. Among 168 PWH less than 50 years old, older age [aOR 1.09 (1.01-1.18)] and no ART use [aOR 40.26 (4.12-393.62)] were associated with hospitalization; age-adjusted comorbidity burden was not ( P  = 0.25). Among 197 PWH at least 50, older age [aOR 1.10 (1.04-1.16)], heart disease [aOR 2.45 (1.04-5.77)], history of HCV [aOR 3.52 (1.29-9.60)], and age-adjusted comorbidity burden [aOR 1.36 (1.12-1.66)] were associated with hospitalization. CONCLUSION: Comorbidity burden is more strongly associated with COVID-19 hospitalization among older, rather than younger, PWH. These findings may have important implications for risk-stratifying COVID-19 therapies and booster recommendations in PWH.

Using Active Surveillance to Identify Monoclonal Antibody Candidates Among COVID-19-Positive Veterans in the Atlanta VA Health Care System.

Background: Strategies for optimizing identification and outreach to potential candidates for monoclonal antibody (Mab) therapy for COVID-19 are not clear. Using a centralized, active surveillance system, the Atlanta Veterans Affairs Health Care System (AVAHCS) infectious disease (ID) team identified candidates for Mab infusion and provided treatment. Observations: As part of a quality improvement project from December 28, 2020, to August 31, 2021, a clinical team consisting of ID pharmacists and physicians reviewed each outpatient with a positive COVID-19 polymerase chain reaction test daily at the AVAHCS. The clinical team used Emergency Use Authorization (EUA) criteria to determine eligibility. Eligible patients were contacted on the same day of review via telephone to confirm eligibility and obtain verbal consent. Telehealth follow-up occurred on day 1 and day 7 postinfusion to assess for adverse events. In total, 2028 patients with COVID-19 were identified; 289 patients (14%) were eligible, and 132 (46%) received Mab therapy. Similar to AVAHCS demographics, a majority of those who received Mab therapy were non-Hispanic Black patients (65%). The most common comorbidities were hypertension (59%) and diabetes (37%). The median time from symptom onset to positive COVID-19 polymerase chain reaction (PCR) test result was 6 days (range, 0-9), and the median time from positive COVID-19 PCR test result to Mab infusion was 2 days (range, 0-8). Twelve patients (9%) required hospitalization for worsening COVID-19 symptoms postinfusion. No deaths occurred. Conclusions: Combining laboratory surveillance and active screening led to high uptake of Mab therapy and minimized delay from symptom onset to Mab infusion, thereby optimizing outpatient treatment of COVID-19. This approach also successfully screened and treated Black patients in the AVAHCS population.

A Challenging Case of Jejunal Dieulafoy's Lesion: A Rare Cause of Refractory Lower-Gastrointestinal Bleeding.

BACKGROUND Dieulafoy's lesion is a rare cause of severe gastrointestinal (GI) bleeding, accounting for approximately 1-2% of all cases of GI hemorrhage. Nevertheless, it can be life-threatening without prompt intervention. Dieulafoy's lesion of jejunal origin can be particularly challenging to identify due to the inability of conventional endoscopic techniques to visualize the jejunum. This case report emphasizes the difficulties in diagnosing and managing jejunal Dieulafoy's lesions and highlights the methods by which to approach refractory bleeding. CASE REPORT This is a case of a 41-year-old man with a history of uncontrolled hypertension who presented with an episode of syncope and melena associated with low hemoglobin levels requiring multiple packed red blood cell transfusions. This warranted searching for a source of bleeding within the gastrointestinal tract via 2 upper-GI endoscopies, a colonoscopy, and an abdominal computed tomography angiogram, all of which failed to localize the site of bleeding. A push enteroscopy was required to identify the lesion in the jejunum, but the bleeding was not controlled despite the application of hemoclips and epinephrine. Consequently, laparotomy and resection of the jejunal segment containing the Dieulafoy's lesion was performed and the diagnosis was established histopathologically. The patient recovered well and was discharged 4 days after the procedure. CONCLUSIONS Suspicion of a jejunal Dieulafoy's lesion should be raised if both upper- and lower-GI endoscopies yield unremarkable findings. Ideally, a push enteroscopy should be utilized diagnostically and to conservatively manage the bleeding. However, laparotomy should be considered in refractory lesions or in the presence of hemodynamic instability.

Clinical Characteristics, Health Care Utilization, and Outcomes Among Patients in a Pilot Surveillance System for Invasive Mold Disease-Georgia, United States, 2017-2019.

Background: Invasive mold diseases (IMDs) cause severe illness, but public health surveillance data are lacking. We describe data collected from a laboratory-based, pilot IMD surveillance system. Methods: During 2017-2019, the Emerging Infections Program conducted active IMD surveillance at 3 Atlanta-area hospitals. We ascertained potential cases by reviewing histopathology, culture, and Aspergillus galactomannan results and classified patients as having an IMD case (based on European Organization for Research and Treatment of Cancer/Invasive Fungal Infections Cooperative Group and the National Institute of Allergy and Infectious Diseases Mycoses Study Group [MSG] criteria) or a non-MSG IMD case (based on the treating clinician's diagnosis and use of mold-active antifungal therapy). We described patient features and compared patients with MSG vs non-MSG IMD cases. Results: Among 304 patients with potential IMD, 104 (34.2%) met an IMD case definition (41 MSG, 63 non-MSG). The most common IMD types were invasive aspergillosis (n = 66 [63.5%]), mucormycosis (n = 8 [7.7%]), and fusariosis (n = 4 [3.8%]); the most frequently affected body sites were pulmonary (n = 66 [63.5%]), otorhinolaryngologic (n = 17 [16.3%]), and cutaneous/deep tissue (n = 9 [8.7%]). Forty-five (43.3%) IMD patients received intensive care unit-level care, and 90-day all-cause mortality was 32.7%; these outcomes did not differ significantly between MSG and non-MSG IMD patients. Conclusions: IMD patients had high mortality rates and a variety of clinical presentations. Comprehensive IMD surveillance is needed to assess emerging trends, and strict application of MSG criteria for surveillance might exclude over one-half of clinically significant IMD cases.

The selenoprotein P 3' untranslated region is an RNA binding protein platform that fine tunes selenocysteine incorporation.

Selenoproteins contain the 21st amino acid, selenocysteine (Sec), which is incorporated at select UGA codons when a specialized hairpin sequence, the Sec insertion sequence (SECIS) element, is present in the 3' UTR. Aside from the SECIS, selenoprotein mRNA 3' UTRs are not conserved between different selenoproteins within a species. In contrast, the 3'-UTR of a given selenoprotein is often conserved across species, which supports the hypothesis that cis-acting elements in the 3'-UTR other than the SECIS exert post-transcriptional control on selenoprotein expression. In order to determine the function of one such SECIS context, we chose to focus on the plasma selenoprotein, SELENOP, which is required to maintain selenium homeostasis as a selenium transport protein that contains 10 Sec residues. It is unique in that its mRNA contains two SECIS elements in the context of a highly conserved 843-nucleotide 3' UTR. Here we have used RNA affinity chromatography and identified PTBP1 as the major RNA binding protein that specifically interacts with the sequence between the two SECIS elements. We then used CRISPR/Cas9 genome editing to delete two regions surrounding the first SECIS element. We found that these sequences are involved in regulating SELENOP mRNA and protein levels, which are inversely altered as a function of selenium concentrations.

The CLIP-170 N-terminal domain binds directly to both F-actin and microtubules in a mutually exclusive manner.

The cooperation between the actin and microtubule (MT) cytoskeletons is important for cellular processes such as cell migration and muscle cell development. However, a full understanding of how this cooperation occurs has yet to be sufficiently developed. The MT plus-end tracking protein CLIP-170 has been implicated in this actin-MT coordination by associating with the actin-binding signaling protein IQGAP1 and by promoting actin polymerization through binding with formins. Thus far, the interactions of CLIP-170 with actin were assumed to be indirect. Here, we demonstrate using high-speed cosedimentation assays that CLIP-170 can bind to filamentous actin (F-actin) directly. We found that the affinity of this binding is relatively weak but strong enough to be significant in the actin-rich cortex, where actin concentrations can be extremely high. Using CLIP-170 fragments and mutants, we show that the direct CLIP-170-F-actin interaction is independent of the FEED domain, the region that mediates formin-dependent actin polymerization, and that the CLIP-170 F-actin-binding region overlaps with the MT-binding region. Consistent with these observations, in vitro competition assays indicate that CLIP-170-F-actin and CLIP-170-MT interactions are mutually exclusive. Taken together, these observations lead us to speculate that direct CLIP-170-F-actin interactions may function to reduce the stability of MTs in actin-rich regions of the cell, as previously proposed for MT end-binding protein 1.

The expression of essential selenoproteins during development requires SECIS-binding protein 2-like.

The dietary requirement for selenium is based on its incorporation into selenoproteins, which contain the amino acid selenocysteine (Sec). The Sec insertion sequence (SECIS) is an RNA structure found in the 3' UTR of all selenoprotein mRNAs, and it is required to convert in-frame UGA codons from termination to Sec-incorporating codons. SECIS-binding protein 2 (Sbp2) is required for Sec incorporation, but its paralogue, SECIS-binding protein 2-like (Secisbp2l), while conserved, has no known function. Here we determined the relative roles of Sbp2 and Secisbp2l by introducing CRISPR mutations in both genes in zebrafish. By monitoring selenoprotein synthesis with 75Se labeling during embryogenesis, we found that sbp2 -/- embryos still make a select subset of selenoproteins but secisbp2l -/- embryos retain the full complement. Abrogation of both genes completely prevents selenoprotein synthesis and juveniles die at 14 days post fertilization. Embryos lacking Sbp2 are sensitive to oxidative stress and express the stress marker Vtg1. We propose a model where Secisbp2l is required to promote essential selenoprotein synthesis when Sbp2 activity is compromised.

Overexpression of the microtubule-binding protein CLIP-170 induces a +TIP network superstructure consistent with a biomolecular condensate.

Proper regulation of microtubule (MT) dynamics is critical for cellular processes including cell division and intracellular transport. Plus-end tracking proteins (+TIPs) dynamically track growing MTs and play a key role in MT regulation. +TIPs participate in a complex web of intra- and inter- molecular interactions known as the +TIP network. Hypotheses addressing the purpose of +TIP:+TIP interactions include relieving +TIP autoinhibition and localizing MT regulators to growing MT ends. In addition, we have proposed that the web of +TIP:+TIP interactions has a physical purpose: creating a dynamic scaffold that constrains the structural fluctuations of the fragile MT tip and thus acts as a polymerization chaperone. Here we examine the possibility that this proposed scaffold is a biomolecular condensate (i.e., liquid droplet). Many animal +TIP network proteins are multivalent and have intrinsically disordered regions, features commonly found in biomolecular condensates. Moreover, previous studies have shown that overexpression of the +TIP CLIP-170 induces large "patch" structures containing CLIP-170 and other +TIPs; we hypothesized that these structures might be biomolecular condensates. To test this hypothesis, we used video microscopy, immunofluorescence staining, and Fluorescence Recovery After Photobleaching (FRAP). Our data show that the CLIP-170-induced patches have hallmarks indicative of a biomolecular condensate, one that contains +TIP proteins and excludes other known condensate markers. Moreover, bioinformatic studies demonstrate that the presence of intrinsically disordered regions is conserved in key +TIPs, implying that these regions are functionally significant. Together, these results indicate that the CLIP-170 induced patches in cells are phase-separated liquid condensates and raise the possibility that the endogenous +TIP network might form a liquid droplet at MT ends or other +TIP locations.