Combinatorial Signal Perception in the BMP Pathway.

The bone morphogenetic protein (BMP) signaling pathway comprises multiple ligands and receptors that interact promiscuously with one another and typically appear in combinations. This feature is often explained in terms of redundancy and regulatory flexibility, but it has remained unclear what signal-processing capabilities it provides. Here, we show that the BMP pathway processes multi-ligand inputs using a specific repertoire of computations, including ratiometric sensing, balance detection, and imbalance detection. These computations operate on the relative levels of different ligands and can arise directly from competitive receptor-ligand interactions. Furthermore, cells can select different computations to perform on the same ligand combination through expression of alternative sets of receptor variants. These results provide a direct signal-processing role for promiscuous receptor-ligand interactions and establish operational principles for quantitatively controlling cells with BMP ligands. Similar principles could apply to other promiscuous signaling pathways.

Dynamics and functional roles of splicing factor autoregulation.

Non-core spliceosome components are essential, conserved regulators of alternative splicing. They provide concentration-dependent control of diverse pre-mRNAs. Many splicing factors direct unproductive splicing of their own pre-mRNAs through negative autoregulation. However, the impact of such feedback loops on splicing dynamics at the single-cell level remains unclear. Here, we developed a system to quantitatively analyze negative autoregulatory splicing dynamics by splicing factor SRSF1 in response to perturbations in single HEK293 cells. We show that negative autoregulatory splicing provides critical functions for gene regulation, establishing a ceiling of SRSF1 protein concentration, reducing cell-cell heterogeneity in SRSF1 levels, and buffering variation in transcription. Most important, it adapts SRSF1 splicing activity to variations in demand from other pre-mRNA substrates. A minimal mathematical model of autoregulatory splicing explains these experimentally observed features and provides values for effective biochemical parameters. These results reveal the unique functional roles that splicing negative autoregulation plays in homeostatically regulating transcriptional programs.

The context-dependent, combinatorial logic of BMP signaling.

Cell-cell communication systems typically comprise families of ligand and receptor variants that function together in combinations. Pathway activation depends on the complex way in which ligands are presented extracellularly and receptors are expressed by the signal-receiving cell. To understand the combinatorial logic of such a system, we systematically measured pairwise bone morphogenetic protein (BMP) ligand interactions in cells with varying receptor expression. Ligands could be classified into equivalence groups based on their profile of positive and negative synergies with other ligands. These groups varied with receptor expression, explaining how ligands can functionally replace each other in one context but not another. Context-dependent combinatorial interactions could be explained by a biochemical model based on the competitive formation of alternative signaling complexes with distinct activities. Together, these results provide insights into the roles of BMP combinations in developmental and therapeutic contexts and establish a framework for analyzing other combinatorial, context-dependent signaling systems.

Ligand-receptor promiscuity enables cellular addressing.

In multicellular organisms, secreted ligands selectively activate, or "address," specific target cell populations to control cell fate decision-making and other processes. Key cell-cell communication pathways use multiple promiscuously interacting ligands and receptors, provoking the question of how addressing specificity can emerge from molecular promiscuity. To investigate this issue, we developed a general mathematical modeling framework based on the bone morphogenetic protein (BMP) pathway architecture. We find that promiscuously interacting ligand-receptor systems allow a small number of ligands, acting in combinations, to address a larger number of individual cell types, defined by their receptor expression profiles. Promiscuous systems outperform seemingly more specific one-to-one signaling architectures in addressing capability. Combinatorial addressing extends to groups of cell types, is robust to receptor expression noise, grows more powerful with increases in the number of receptor variants, and is maximized by specific biochemical parameter relationships. Together, these results identify design principles governing cellular addressing by ligand combinations.

Predictors of humoral response to SARS-CoV-2 mRNA vaccine BNT162b2 in patients receiving maintenance dialysis.

Objective: Patients on dialysis are at high risk for severe COVID-19 and associated morbidity and mortality. We examined the humoral response to SARS-CoV-2 mRNA vaccine BNT162b2 in a maintenance dialysis population. Design: Single-center cohort study. Setting and participants: Adult maintenance dialysis patients at 3 outpatient dialysis units of a large academic center. Methods: Participants were vaccinated with 2 doses of BNT162b2, 3 weeks apart. We assessed anti-SARS-CoV-2 spike antibodies (anti-S) ∼4-7 weeks after the second dose and evaluated risk factors associated with insufficient response. Definitions of antibody response are as follows: nonresponse (anti-S level, <50 AU/mL), low response (anti-S level, 50-839 AU/mL), and sufficient response (anti-S level, ≥840 AU/mL). Results: Among the 173 participants who received 2 vaccine doses, the median age was 60 years (range, 28-88), 53.2% were men, 85% were of Black race, 86% were on in-center hemodialysis and 14% were on peritoneal dialysis. Also, 7 participants (4%) had no response, 27 (15.6%) had a low response, and 139 (80.3%) had a sufficient antibody response. In multivariable analysis, factors significantly associated with insufficient antibody response included end-stage renal disease comorbidity index score ≥5 and absence of prior hepatitis B vaccination response. Conclusions: Although most of our study participants seroconverted after 2 doses of BNT162b2, 20% of our cohort did not achieve sufficient humoral response. Our findings demonstrate the urgent need for a more effective vaccine strategy in this high-risk patient population and highlight the importance of ongoing preventative measures until protective immunity is achieved.

Accessible Medical Education & TIC: Increasing Equitable Care for Disabled Patients.

An estimated 1 in 4 U.S. adults has a disability, and this number continues to increase. Disabled individuals face significant healthcare inequities, including but not limited to inaccessibility and mistreatment. Our current healthcare system is ill-equipped to provide equitable care to this population. There is a lack of accessibility in healthcare environments, lack of accessible medical training to enable disabled people to become healthcare providers serving their own community, and lack of thorough medical education that encompasses care for disabled patients. Furthermore, the increased risk of trauma, as well as increased risk of medical trauma specifically, endured by disabled people puts them at greater risk of long-lasting adverse effects. In this commentary, we analyze three key areas: 1) the current state of healthcare for disabled patients, 2) disability in medical education & physician workforce, and 3) the relationship between trauma and disability. We argue that the road to more equitable care for disabled patients involves changes to medical education that address all three of these areas. Medical training should expose trainees to disability early and throughout their training, should be made more accessible to support disabled physicians, and finally, should be trauma-informed in a manner that explicitly includes caring for disabled patients and their other intersecting identities.

Two-Color Imaging of Nonrepetitive Endogenous Loci in Human Cells.

Tools for live cell imaging of multiple nonrepetitive genomic loci in mammalian cells are necessary to study chromatin dynamics. Here, we report a new system based on two chromosomally integrated orthogonal irregular repeat arrays and particularly a new general strategy to construct irregular repeat arrays. Briefly, we utilized a "bridge oligonucleotide-mediated ligation" protocol to assemble 8-mer repeats de novo which were then combined into a final 96-mer repeat array using Golden Gate cloning. This strategy was used for assembling a new mutant TetO irregular repeat array, which worked orthogonally to the wild type TetO repeat. Single copy integration of the new repeat array did not cause replication deficiencies at the tagged locus. Moreover, the mutant TetO irregular repeat could also be visualized by CRISPR imaging. Our new irregular repeat assembly method demonstrates a generally applicable strategy that can be used for assembling additional orthogonal repeat arrays for imaging genomic loci and irregular repeats to visualize RNA or proteins via signal amplification.

Atypical femur fractures: 81 individual personal histories.

CONTEXT: An online voluntary association of individuals who had incurred one or more atypical femur fractures (AFFs) while taking bisphosphonates for prevention or treatment of osteoporosis provided an opportunity to collect long-term histories. SETTING: Individuals from a nationwide general community completed an anonymous survey documenting their history. PARTICIPANTS: Within a larger cadre, cases were selected where the documentation, including fracture radiographs, verified the diagnosis based on published standards. Eighty-one of this group responded to the anonymous survey. INTERVENTIONS: We describe passively collected observational data only. OUTCOME MEASURES: The incidence of a large number of potential variations was determined. RESULTS: The mean duration of bisphosphonate treatment was 9.5 yr. Prevention was the initial indication in 68% of the subjects; 94% started on alendronate, 77% reported prodromal pain, only 16% of these were diagnosed with incident stress fractures, and 39.5% experienced a contralateral AFF from less than 1 month to 49 months after the first. Of 71 subjects with a completed first AFF, 38% reported delayed healing, 11% had a complete contralateral AFF, and 22% underwent prophylactic rodding for a contralateral stress AFF. Forty-four percent of subjects with complete AFFs were continued on a bisphosphonate after the fracture. Thirty-five percent incurred a metatarsal fracture. CONCLUSIONS: AFF patients experienced delayed healing, prodromal pain, and persisting risk of a contralateral and/or other fracture. Femur pain evaluation in patients on long-term bisphosphonates may facilitate early diagnosis of stress AFFs, permitting intervention, thus reducing completed and/or contralateral or other fracture risk. The details of these histories will assist in counseling regarding prognosis after an initial AFF.