Calpain activity is negatively regulated by a KCTD7-Cullin-3 complex via non-degradative ubiquitination

Calpains are a class of non-lysosomal cysteine proteases that exert their regulatory functions via limited proteolysis of their substrates. Similar to the lysosomal and proteasomal systems, calpain dysregulation is implicated in the pathogenesis of neurodegenerative disease and cancer. Despite intensive efforts placed on the identification of mechanisms that regulate calpains, however, calpain protein modifications that regulate calpain activity are incompletely understood. Here we show that calpains are regulated by KCTD7, a cytosolic protein of previously uncharacterized function whose pathogenic mutations result in epilepsy, progressive ataxia, and severe neurocognitive deterioration. We show that KCTD7 works in complex with Cullin-3 and Rbx1 to execute atypical, non-degradative ubiquitination of calpains at specific sites (K398 of calpain 1, and K280 and K674 of calpain 2). Experiments based on single-lysine mutants of ubiquitin determined that KCTD7 mediates ubiquitination of calpain 1 via K6-, K27-, K29-, and K63-linked chains, whereas it uses K6-mediated ubiquitination to modify calpain 2. Loss of KCTD7-mediated ubiquitination of calpains led to calpain hyperactivation, aberrant cleavage of downstream targets, and caspase-3 activation. CRISPR/Cas9-mediated knockout of Kctd7 in mice phenotypically recapitulated human KCTD7 deficiency and resulted in calpain hyperactivation, behavioral impairments, and neurodegeneration. These phenotypes were largely prevented by pharmacological inhibition of calpains, thus demonstrating a major role of calpain dysregulation in KCTD7-associated disease. Finally, we determined that Cullin-3-KCTD7 mediates ubiquitination of all ubiquitous calpains. These results unveil a novel mechanism and potential target to restrain calpain activity in human disease and shed light on the molecular pathogenesis of KCTD7-associated disease

Factors Associated With COVID-19 Death in the United States: Cohort Study

BackgroundSince the initial COVID-19 cases were identified in the United States in February 2020, the United States has experienced a high incidence of the disease. Understanding the risk factors for severe outcomes identifies the most vulnerable populations and helps in decision-making. ObjectiveThis study aims to assess the factors associated with COVID-19–related deaths from a large, national, individual-level data set. MethodsA cohort study was conducted using data from the Optum de-identified COVID-19 electronic health record (EHR) data set; 1,271,033 adult participants were observed from February 1, 2020, to August 31, 2020, until their deaths due to COVID-19, deaths due to other reasons, or the end of the study. Cox proportional hazards models were constructed to evaluate the risks for each patient characteristic. ResultsA total of 1,271,033 participants (age: mean 52.6, SD 17.9 years; male: 507,574/1,271,033, 39.93%) were included in the study, and 3315 (0.26%) deaths were attributed to COVID-19. Factors associated with COVID-19–related death included older age (≥80 vs 50-59 years old: hazard ratio [HR] 13.28, 95% CI 11.46-15.39), male sex (HR 1.68, 95% CI 1.57-1.80), obesity (BMI ≥40 vs <30 kg/m2: HR 1.71, 95% CI 1.50-1.96), race (Hispanic White, African American, Asian vs non-Hispanic White: HR 2.46, 95% CI 2.01-3.02; HR 2.27, 95% CI 2.06-2.50; HR 2.06, 95% CI 1.65-2.57), region (South, Northeast, Midwest vs West: HR 1.62, 95% CI 1.33-1.98; HR 2.50, 95% CI 2.06-3.03; HR 1.35, 95% CI 1.11-1.64), chronic respiratory disease (HR 1.21, 95% CI 1.12-1.32), cardiac disease (HR 1.10, 95% CI 1.01-1.19), diabetes (HR 1.92, 95% CI 1.75-2.10), recent diagnosis of lung cancer (HR 1.70, 95% CI 1.14-2.55), severely reduced kidney function (HR 1.92, 95% CI 1.69-2.19), stroke or dementia (HR 1.25, 95% CI 1.15-1.36), other neurological diseases (HR 1.77, 95% CI 1.59-1.98), organ transplant (HR 1.35, 95% CI 1.09-1.67), and other immunosuppressive conditions (HR 1.21, 95% CI 1.01-1.46). ConclusionsThis is one of the largest national cohort studies in the United States; we identified several patient characteristics associated with COVID-19–related deaths, and the results can serve as the basis for policy making. The study also offered directions for future studies, including the effect of other socioeconomic factors on the increased risk for minority groups

Additional file 1 of Defining critical illness using immunological endotypes in patients with and without sepsis: a cohort study

Additional file 1. Supplemental Materials: Methods