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We are 52 Black scientists. Here, we establish the context of Juneteenth in STEMM and discuss the barriers Black scientists face, the struggles they endure, and the lack of recognition they receive. We review racism's history in science and provide institutional-level solutions to reduce the burdens on Black scientists.

Protein aggregates are a common feature of diseased and aged cells. Membrane proteins comprise a quarter of the proteome, and yet, it is not well understood how aggregation of membrane proteins is regulated and what effects these aggregates can have on cellular health. We have determined in yeast that the derlin Dfm1 has a chaperone-like activity that influences misfolded membrane protein aggregation. We establish that this function of Dfm1 does not require recruitment of the ATPase Cdc48 and it is distinct from Dfm1's previously identified function in dislocating misfolded membrane proteins from the endoplasmic reticulum (ER) to the cytosol for degradation. Additionally, we assess the cellular impacts of misfolded membrane proteins in the absence of Dfm1 and determine that misfolded membrane proteins are toxic to cells in the absence of Dfm1 and cause disruptions to proteasomal and ubiquitin homeostasis.

Nearly one-third of proteins are initially targeted to the endoplasmic reticulum (ER) membrane, where they are correctly folded and then delivered to their final cellular destinations. To prevent the accumulation of misfolded membrane proteins, ER-associated degradation (ERAD) moves these clients from the ER membrane to the cytosol, a process known as retrotranslocation. Our recent work in Saccharomyces cerevisiae reveals a derlin rhomboid pseudoprotease, Dfm1, is involved in the retrotranslocation of ubiquitinated ERAD membrane substrates. In this study, we identify conserved residues of Dfm1 that are critical for retrotranslocation. We find several retrotranslocation-deficient Loop 1 mutants that display impaired binding to membrane substrates. Furthermore, Dfm1 possesses lipid thinning function to facilitate in the removal of ER membrane substrates, and this feature is conserved in its human homolog, Derlin-1, further implicating that derlin-mediated retrotranslocation is a well-conserved process.

Cells are equipped with protein quality control pathways in order to maintain a healthy proteome; a process known as protein homeostasis. Dysfunction in protein homeostasis leads to the development of many diseases that are associated with proteinopathies. Recently, the rhomboid superfamily has attracted much attention concerning their involvement in protein homeostasis. While their functional role has become much clearer in the last few years, their systemic significance in mammals remains elusive. Here we delineate the current knowledge of rhomboids in protein quality control and how these functions are integrated at the organismal level.

Prokaryotes have aerobic and anaerobic electron acceptors for oxidative folding of periplasmic proteins. The mitochondrial intermembrane space has an analogous pathway with the oxidoreductase Mia40 and sulfhydryl oxidase Erv1, termed the mitochondrial intermembrane space assembly (MIA) pathway. The aerobic electron acceptors include oxygen and cytochrome c, but an acceptor that can function under anaerobic conditions has not been identified. Here we show that the fumarate reductase Osm1, which facilitates electron transfer from fumarate to succinate, fills this gap as a new electron acceptor. In addition to microsomes, Osm1 localizes to the mitochondrial intermembrane space and assembles with Erv1 in a complex. In reconstitution studies with reduced Tim13, Mia40, and Erv1, the addition of Osm1 and fumarate completes the disulfide exchange pathway that results in Tim13 oxidation. From in vitro import assays, mitochondria lacking Osm1 display decreased import of MIA substrates, Cmc1 and Tim10. Comparative reconstitution assays support that the Osm1/fumarate couple accepts electrons with similar efficiency to cytochrome c and that the cell has strategies to coordinate expression of the terminal electron acceptors. Thus Osm1/fumarate is a new electron acceptor couple in the mitochondrial intermembrane space that seems to function in both aerobic and anaerobic conditions.

In the face of a challenging climate STEM (Science, Technology, Engineering and Mathematics) higher education that is resistant to Diversity, Equity, and Inclusion efforts aimed to increase and retain students from historically excluded groups (HEGs), there is a critical need for a support structure to ensure students from HEGs continue to be recruited retained. The Biology Undergraduate and Master's Mentorship Program (BUMMP) embodies this commitment to fostering scientific identity, efficacy, and a sense of belonging for first‐generation and historically underserved undergraduate and master's students at UC San Diego. The mission of BUMMP is to cultivate a sense of belonging, instill confidence, and nurture a strong scientific identity amongst all its participants. At its core, the three pillars of BUMMP are (1) mentorship, (2) professional development, and (3) research. Quality mentorship is provided where students receive personal guidance from faculty, graduate students, postdocs, and industry leaders in navigating their career pathways. Complementing mentorship, BUMMP provides paid research opportunities and prioritizes professional development by offering workshops designed to enhance students' professional skills. These three pillars form the backbone of BUMMP, empowering students from all backgrounds and ensuring their retention and persistence in STEM. So far, we've served over 1350 mentees, collaborated with 809 mentors, and had over 180 mentees actively engaged in BUMMP‐sponsored research activities. The primary focus of this paper is to provide a programmatic guideline for the three pillars of BUMMP: mentorship, professional development, and research. This will offer a blueprint for other institutions to establish similar mentorship programs. Additionally, the paper highlights the impact of the BUMMP program and surveyed mentees who have participated in the mentorship and research component of BUMMP. We showed that mentorship and research experience enhance students' sense of belonging, science identity, and science efficacy, which are key predictors of retention and persistence in pursuing a STEM career. Overall, BUMMP's expansive efforts have made a tremendous impact at UC San Diego and will continue to foster a community of future leaders who will be prepared to make meaningful contributions to the scientific community and beyond. [ABSTRACT FROM AUTHOR]

The HRD (HMG-CoA reductase degradation) pathway is a conserved route of endoplasmic reticulum-associated degradation (ERAD), by which misfolded ER proteins are ubiquitinated and degraded. ERAD substrates are ubiquitinated by the action of the Hrd1 RING-H2 E3 ligase. Hrd1 is always present in a stoichiometric complex with the ER membrane protein Hrd3, which is also required for HRD-dependent degradation. Despite its conserved presence, unequivocal study of Hrd3 function has been precluded by its central role in Hrd1 stability. Loss of Hrd3 causes unrestricted self-degradation of Hrd1, resulting in significant loss of the core ligase. Accordingly, the degree to which Hrd3 functions independently of Hrd1 stabilization has remained unresolved. By capitalizing on our studies of Usa1 in Hrd1 degradation, we have devised a new approach to evaluate Hrd3 functions in ERAD. We now show that Hrd3 has a direct and critical role in ERAD in addition to Hrd1 stabilization. This direct component of Hrd3 is phenotypically as important as Hrd1 in the native HRD complex. Hrd3 was required the E3 activity of Hrd1, rather than substrate or E2 recruitment to Hrd1. Although Hrd1 can function in some circumstances independent of Hrd3, these studies show an indispensable role for Hrd3 in living cells.

A redox-regulated import pathway consisting of Mia40 and Erv1 mediates the import of cysteine-rich proteins into the mitochondrial intermembrane space. Mia40 is the oxidoreductase that inserts two disulfide bonds into the substrate simultaneously. However, Mia40 has one redox-active cysteine pair, resulting in ambiguity about how Mia40 accepts numerous electrons during substrate oxidation. In this study, we have addressed the oxidation of Tim13 in vitro and in organello. Reductants such as glutathione and ascorbate inhibited both the oxidation of the substrate Tim13 in vitro and the import of Tim13 and Cmc1 into isolated mitochondria. In addition, a ternary complex consisting of Erv1, Mia40, and substrate, linked by disulfide bonds, was not detected in vitro. Instead, Mia40 accepted six electrons from substrates, and this fully reduced Mia40 was sensitive to protease, indicative of conformational changes in the structure. Mia40 in mitochondria from the erv1-101 mutant was also trapped in a completely reduced state, demonstrating that Mia40 can accept up to six electrons as substrates are imported. Therefore, these studies support that Mia40 functions as an electron sink to facilitate the insertion of two disulfide bonds into substrates.

Nearly one‐third of nascent proteins are initially targeted to the endoplasmic reticulum (ER), where they are correctly folded and assembled before being delivered to their final cellular destinations. To prevent the accumulation of misfolded membrane proteins, ER‐associated degradation (ERAD) removes these client proteins from the ER membrane to the cytosol in a process known as retrotranslocation. Our previous work demonstrated that rhomboid pseudoprotease Dfm1 is involved in the retrotranslocation of ubiquitinated membrane integral ERAD substrates. Herein, we found that Dfm1 associates with the SPOTS complex, which is composed of serine palmitoyltransferase (SPT) enzymes and accessory components that are critical for catalyzing the first rate‐limiting step of the sphingolipid biosynthesis pathway. Furthermore, Dfm1 employs an ERAD‐independent role for facilitating the ER export and endosome‐ and Golgi‐associated degradation (EGAD) of Orm2, which is a major antagonist of SPT activity. Given that the accumulation of human Orm2 homologs, ORMDLs, is associated with various pathologies, our study serves as a molecular foothold for understanding how dysregulation of sphingolipid metabolism leads to various diseases. Synopsis: Previously, yeast derlin Dfm1 was found to be critical for retrotranslocating integral membrane substrates targeted for ER‐associated degradation (ERAD). This study reveals an ERAD‐independent role for Dfm1 in regulating the degradation of the negative regulator of sphingolipid biosynthesis, Orm2. Dfm1 deletion leads to dysregulated levels of early precursors of the sphingolipid biosynthesis pathway.Derlin Dfm1 associates with rate‐limiting enzymes and regulators of the sphingolipid biosynthesis pathway.Dfm1 targets Ypk1‐dependent phosphorylated Orm2 for ER export and endosome‐ and Golgi‐associated degradation (EGAD).Dfm1's Cdc48‐recruitment function is not required for regulating Orm2 degradation. [ABSTRACT FROM AUTHOR]

Cells are equipped with protein quality control pathways in order to maintain a healthy proteome; a process known as protein homeostasis. Dysfunction in protein homeostasis leads to the development of many diseases that are associated with proteinopathies. Recently, the rhomboid superfamily has attracted much attention concerning their involvement in protein homeostasis. While their functional role has become much clearer in the last few years, their systemic significance in mammals remains elusive. Here we delineate the current knowledge of rhomboids in protein quality control and how these functions are integrated at the organismal level. [ABSTRACT FROM AUTHOR]

Rhomboid proteases are ubiquitous intramembrane serine proteases that can cleave transmembrane substrates within lipid bilayers. They exhibit many and diverse functions, such as but not limited to, growth factor signaling, immune and inflammatory response, protein quality control, and parasitic invasion. Human rhomboid protease RHBDL4 has been demonstrated to play a critical role in removing misfolded proteins from the Endoplasmic Reticulum and is implicated in severe diseases such as various cancers and Alzheimer's disease. Therefore, RHBDL4 is expected to constitute an important therapeutic target for such devastating diseases. Despite its critical role in many biological processes, the enzymatic properties of RHBDL4 remain largely unknown. To enable a comprehensive characterization of RHBDL4's kinetics, catalytic parameters, substrate specificity, and binding modality we expressed and purified recombinant RHBDL4, and employed it in a Förster Resonance Energy Transfer-based cleavage assay. Until now, kinetic studies have been limited mostly to bacterial rhomboid proteases. Our in vitro platform offers a new method for studying RHBDL4's enzymatic function and substrate preferences. Furthermore, we developed and tested potential inhibitors using our assay and successfully identified peptidyl α-ketoamide inhibitors of RHBDL4 that are highly effective against recombinant RHBDL4. We utilize ensemble docking and molecular dynamics (MD) simulations to explore the binding modality of substrate-derived peptides bound to RHBDL4. Our analysis focused on key interactions and dynamic movements within RHBDL4's active site that contributed to binding stability, offering valuable insights for optimizing the non-prime side of RHBDL4 ketoamide inhibitors. In summary, our study offers fundamental insights into RHBDL4's catalytic activities and substrate preferences, laying the foundation for downstream applications such as drug inhibitor screenings and structure-function studies, which will enable the identification of lead drug compounds for RHBDL4.

The endoplasmic reticulum (ER) is an essential organelle in eukaryotic cells and is a major site for protein folding, modification, and lipid synthesis. Perturbations within the ER, such as protein misfolding and high demand for protein folding, lead to dysregulation of the ER protein quality control network and ER stress. Recently, the rhomboid superfamily has emerged as a critical player in ER protein quality control because it has diverse cellular functions, including ER-associated degradation (ERAD), endosome Golgi-associated degradation (EGAD), and ER preemptive quality control (ERpQC). This breadth of function both illustrates the importance of the rhomboid superfamily in health and diseases and emphasizes the necessity of understanding their mechanisms of action. Because dysregulation of rhomboid proteins has been implicated in various diseases, such as neurological disorders and cancers, they represent promising potential therapeutic drug targets. This review provides a comprehensive account of the various roles of rhomboid proteins in the context of ER protein quality control and discusses their significance in health and disease., (Copyright © 2023 Cold Spring Harbor Laboratory Press; all rights reserved.)