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Abstract Proximity ligation assays (PLAs) use specific antibodies to detect endogenous protein‐protein interactions. PLAs are a highly useful biochemical technique that allow two proteins within proximity to be visualized with fluorescent probes amplified by PCR. While this technique has gained prominence, the use of a PLA in mouse skeletal muscle (SkM) is novel. In this article, we discuss how the PLA method can be used in SkM to study the protein‐protein interactions within mitochondria‐endoplasmic reticulum contact sites (MERCs). © 2025 The Author(s). Current Protocols published by Wiley Periodicals LLC. Basic Protocol: Proximity ligation assay for skeletal muscle tissue and myoblast for MERC proteins 

Abstract OPA1 is a dynamin‐related GTPase that modulates mitochondrial dynamics and cristae integrity. Humans carry eight different isoforms of OPA1 and mice carry five, all of which are expressed as short‐ or long‐form isoforms. These isoforms contribute to OPA1's ability to control mitochondrial energetics and DNA maintenance. However, western blot isolation of all long and short isoforms of OPA1 can be difficult. To address this issue, we developed an optimized western blot protocol based on improving running time to isolate five different isoforms of OPA1 in mouse cells and tissues. This protocol can be applied to study changes in mitochondrial structure and function. © 2025 The Author(s). Current Protocols published by Wiley Periodicals LLC. Basic Protocol: Western Blot Protocol for Isolating OPA1 Isoforms in Mouse Primary Skeletal Muscle Cells 

Tweetable abstract Mitochondrial transplantation has been used to treat various diseases associated with mitochondrial dysfunction. Here, we highlight the considerations in quality control mechanisms that should be considered in the context of mitochondrial transplantation. 

Tweetable abstract This perspective considers several avenues for future research on mitochondrial dynamics, stress, and DNA in outer space. 

Abstract The physical characteristics of brown adipose tissue (BAT) are defined by the presence of multilocular lipid droplets (LDs) within the brown adipocytes and a high abundance of iron‐containing mitochondria, which give it its characteristic color. Normal mitochondrial function is, in part, regulated by organelle‐to‐organelle contacts. For example, the contact sites that mediate mitochondria–LD interactions are thought to have various physiological roles, such as the synthesis and metabolism of lipids. Aging is associated with mitochondrial dysfunction, and previous studies show that there are changes in mitochondrial structure and the proteins that modulate organelle contact sites. However, how mitochondria–LD interactions change with aging has yet to be fully clarified. Therefore, we sought to define age‐related changes in LD morphology and mitochondria–lipid interactions in BAT. We examined the three‐dimensional morphology of mitochondria and LDs in young (3‐month) and aged (2‐year) murine BAT using serial block face‐scanning electron microscopy and the Amira program for segmentation, analysis, and quantification. Our analyses showed reductions in LD volume, area, and perimeter in aged samples in comparison to young samples. Additionally, we observed changes in LD appearance and type in aged samples compared to young samples. Notably, we found differences in mitochondrial interactions with LDs, which could implicate that these contacts may be important for energetics in aging. Upon further investigation, we also found changes in mitochondrial and cristae structure for the mitochondria interacting with LDs. Overall, these data define the nature of LD morphology and organelle–organelle contacts during aging and provide insight into LD contact site changes that interconnect biogerontology with mitochondrial function, metabolism, and bioactivity in aged BAT. 

Abstract A first‐generation college student is typically defined as a student whose biological parent(s) or guardian(s) never attended college or who started but did not finish college. However, “first‐generation” can represent diverse family education situations. The first‐generation student community is a multifaceted, and intersectional group of individuals who frequently lack educational/financial resources to succeed and, consequently, require supportive environments with rigorous mentorship. However, first‐generation students often do not make their identity as first‐generation students known to others due to several psychosocial and academic factors. Therefore, they are often “invisible minorities” in higher education. In this paper, we describe the diverse family situations of first‐generation students, further define “first‐generation,” and suggest five actions that first‐generation trainees at the undergraduate/graduate stages can engage in to succeed in an academic climate. We also provide suggestions for mentors to accommodate first‐generation students' unique experiences and equip them with tools to deliver intentional mentoring practices. We hope that this paper will help promote first‐generation student success throughout the academic pipeline. 

Abstract Machine learning has proven useful in analyzing complex biological data and has greatly influenced the course of research in structural biology and precision medicine. Deep neural network models oftentimes fail to predict the structure of complex proteins and are heavily dependent on experimentally determined structures for their training and validation. Single‐particle cryogenic electron microscopy (cryoEM) is also advancing the understanding of biology and will be needed to complement these models by continuously supplying high‐quality experimentally validated structures for improvements in prediction quality. In this perspective, the significance of structure prediction methods is highlighted, but the authors also ask, what if these programs cannot accurately predict a protein structure important for preventing disease? The role of cryoEM is discussed to help fill the gaps left by artificial intelligence predictive models in resolving targetable proteins and protein complexes that will pave the way for personalized therapeutics.