Biology subjects

Mills, J. C.

Publications and source records attributed to Mills, J. C..

9 recordsLinked to original sources

bioRxiv2026

Stress-Responsive Protein IFRD1 Protects Assembled Ribosomes via a Ribosome-Salvaging Mechanism

The ability of epithelial cells to cope with injury and undergo regeneration depends on tightly coordinated cellular responses. IFRD1 is a stress-responsive protein that is evolutionarily conserved and required for the cellular regeneration program paligenosis; however, how IFRD1 works in paligenosis is not known. Here we demonstrate that IFRD1 is primarily a cytosolic ribosome-binding protein, specifically binding 80S monosomes that are not actively engaged in translation. Using multiple in vivo and in vitro injury models, including cerulein-induced pancreatitis in mice and tunicamycin-induced ER stress in cell culture, we demonstrate that IFRD1 acts as a ribosome-salvaging factor, preventing ribosomes from degradation. In the absence of IFRD1 during ER stress, non-translating 80S ribosomes were unstable and prone to disassembly and selective degradation. The resulting accumulation of degraded ribosomal subunits overwhelmed cellular autophagic machinery, as evidenced by accumulation of the autophagy-tagging protein p62, even though overall autophagic flux remained unaffected. Ultimately, cells lacking IFRD1 showed reduced mTORC1 activity followed by increased cell death, consistent with patterns observed in cells lacking IFRD1 during paligenosis. Thus, we detail a previously unrecognized cellular function for IFRD1 in stabilizing and preserving the mature ribosome pool during metabolic and translational transitions such as paligenosis.

cell biology↗

bioRxiv2026

Evidence that injury can cause Drosophila gut differentiated, polyploid enterocytes to be recruited as stem cells via paligenosis

Differentiated cells can return to a progenitor-like state in response to injury via the evolutionarily conserved cellular program called paligenosis. Paligenosis proceeds by three stages: 1) autophagy/autodegradation of differentiated cell architecture, 2) metaplasia/progenitor gene induction, 3) TOR complex 1 (TORC1)-dependent cell cycle re-entry. Using multiple injury and reverse-lineage-tracing approaches in the Drosophila gut, we show that mature polyploid enterocytes dedifferentiate into diploid progenitors in response to epithelial injury. Several key findings suggest a role for paligenosis. Shortly after injury, enterocytes dramatically increased autophagic flux (stage 1); additionally, pharmacological and genetic inhibition of autophagy blocked progenitor recruitment. Rapamycin also blocked recruitment, indicating that TORC1 is required (stage 3). Finally, RNAi knockdown of ifrd1, an evolutionarily conserved protein required for paligenosis, blocked progenitor recruitment. Thus, replenishment of diploid progenitors from differentiated polyploid cells may occur by paligenosis. The Drosophila gut may offer a versatile system for dissecting the mechanisms of this evolutionarily conserved pathway. Impact StatementMature, polyploid Drosophila enterocytes may dedifferentiate to a stem cell-like, diploid state via the paligenosis cell regeneration program, adding to evidence that paligenosis is a fundamental cellular process and highlighting Drosophila gut as a potential model system for its study.

developmental biology↗

bioRxiv2025

Autophagy-Dependent Regulation of YAP1 by STK38 Governs Recruitment of Differentiated Cells as Progenitor Cells During Regeneration

Paligenosis is a conserved cellular plasticity program that allows mature cells to reenter the cell cycle in response to tissue injury. Paligenosis progresses via three stages: autodegradation (with dramatic increase in autophagy and lysosomes), induction of metaplastic or fetal-like genes, and cell cycle entry. Hippo signaling, particularly the downstream effector YAP1, regulates cellular plasticity, but its role in paligenosis has not been studied. Here we first examine paligenosis in digestive-enzyme-secreting chief cells in mouse stomach. We identify Serine/Threonine Kinase 38 (STK38) as a non-canonical YAP1 kinase that phosphorylated and deactivated YAP1 in uninjured chief cells. During paligenosis, STK38 was degraded by autophagy in stage 1, dephosphorylating and activating YAP1. YAP1 activation was necessary and sufficient for the paligenosis that converts chief cells into metaplastic, proliferating progenitors. Additionally, we show STK38, like canonical Hippo kinases, interact with NF2. We also observed the same pattern of YAP1 induction via autophagic destruction of STK38 in other tissues and cell types, suggesting a universal logic model for how the massive autophagy activated in differentiated cells during tissue damage can consequently activate Hippo effectors to induce plasticity for tissue regeneration.

developmental biology↗

bioRxiv2024

Inhibition of Ribosome Biogenesis in vivo Causes p53-Dependent Death and p53-Independent Dysfunction

Ribosomes are critical for cell function; their synthesis (known as ribosome biogenesis; "RiBi") is complex and energy-intensive. Surprisingly little is known about RiBi in differentiated cells in vivo in adult tissue. Here, we generated mice with conditional deletion of Nat10, an essential gene for RiBi and translation, to investigate effects of RiBi blockade in vivo. We focused on RiBi in a long-lived, ribosome-rich cell population, pancreatic acinar cells, during homeostasis and tumorigenesis. We observed a surprising latency of several weeks between Nat10 deletion and onset of structural and functional abnormalities and p53-dependent acinar cell death, which was associated with translocation of ribosomal proteins RPL5 and RPL11 into acinar cell nucleoplasm. Indeed, deletion of Trp53 could rescue acinar cells from apoptotic cell death; however, Nat10{Delta}/{Delta}; Trp53{Delta}/{Delta} acinar cells remained morphologically and functionally abnormal. Moreover, the deletion of Trp53 did not rescue the lethality of inducible, globally deleted Nat10 in adult mice nor did it rescue embryonic lethality of global Nat10 deletion, emphasizing p53-independent consequences of RiBi inhibition. Deletion of Nat10 in acinar cells blocked Kras-oncogene-driven pancreatic intraepithelial neoplasia and subsequent pancreatic ductal adenocarcinoma, regardless of Trp53 mutation status. Together, our results provide initial insights into how cells respond to defects in RiBi and translation in vivo.

cell biology↗

bioRxiv2024

Cathartocytosis: How Cells Jettison Unwanted Material as They Reprogram

Injury can cause differentiated cells to undergo massive reprogramming to become proliferative to repair tissue via a cellular program called paligenosis. Gastric digestive-enzyme-secreting chief cells use paligenosis to reprogram into progenitor-like Spasmolytic-Polypeptide Expressing Metaplasia (SPEM) cells. Stage 1 of paligenosis is the downscaling of mature cell architecture via a process involving lysosomes. Here, we noticed that sulfated glycoproteins were not only digested during paligenosis but also excreted into the gland lumen. Various genetic and pharmacological approaches showed that endoplasmic reticulum membranes and secretory granule cargo were also excreted and that the process proceeded in parallel with, but was mechanistically independent of autophagy. 3-dimensional light and electron-microscopy demonstrated that excretion occurred via unique, complex, multi-chambered invaginations of the apical plasma membrane. As this lysosome-independent cell cleansing process does not seem to have been priorly described, we termed it "cathartocytosis". Cathartocytosis allows a cell to rapidly eject excess material without waiting for autophagic and lysosomal digestion. We speculate the ejection of sulfated glycoproteins would aid in downscaling and might also help bind and flush pathogens away from tissue.

cell biology↗

bioRxiv2024

A new role for IFRD1 in regulation of ER stress in bladder epithelial homeostasis

A healthy bladder requires the homeostatic maintenance of and rapid regeneration of urothelium upon stress/injury/infection. Several factors have been identified to play important roles in urothelial development, injury and disease response, however, little is known about urothelial regulation at homeostasis. Here, we identify a new role for IFRD1, a stress-induced gene that has recently been demonstrated to play a critical role in adult tissue proliferation and regeneration, in maintenance of urothelial function/ homeostasis in a mouse model. We show that the mouse bladder expresses IFRD1 at homeostasis and its loss alters the global transcriptome of the bladder with significant accumulation of cellular organelles including multivesicular bodies with undigested cargo, lysosomes and mitochondria. We demonstrate that IFRD1 interacts with several mRNA-translation-regulating factors in human urothelial cells and that the urothelium of Ifrd1-/- mice reveal decreased global translation and enhanced endoplasmic reticulum (ER) stress response. Ifrd1-/- bladders have activation of the unfolded protein response (UPR) pathway, specifically the PERK arm, with a concomitant increase in oxidative stress and spontaneous exfoliation of urothelial cells. Further, we show that such increase in cell shedding is associated with a compensatory proliferation of the basal cells but impaired regeneration of superficial cells. Finally, we show that upon loss of IFRD1, mice display aberrant voiding behavior. Thus, we propose that IFRD1 is at the center of many crucial cellular pathways that work together to maintain urothelial homeostasis, highlighting its importance as a target for diagnosis and/or therapy in bladder conditions.

cell biology↗

bioRxiv2022

Pancreas resident macrophage-induced fibrosis has divergent roles in pancreas inflammatory injury and PDAC

Tissue-resident macrophages (TRMs) are long-lived cells that maintain locally and can be phenotypically distinct from monocyte-derived macrophages (MDMs). However, whether TRMs and MDMs have functional distinction under differing pathologies is not understood. Here, we show a significant portion of macrophages that accumulated during pancreatitis and pancreatic cancer were expanded from TRMs. We further established that pancreas TRMs have a distinct extracellular matrix remodeling phenotype that was critical for maintaining tissue homeostasis during inflammation. Loss of TRMs led to exacerbation of severe pancreatitis and animal death, due to impaired acinar cell survival and recovery. In pancreatitis, TRMs elicited protective effects by triggering the accumulation and activation of fibroblasts, which was necessary for initiating fibrosis as a wound healing response. The same TRM-driven fibrosis, however, drove pancreas cancer pathogenesis and progression. Together, these findings indicate that TRMs play divergent roles in the pathogenesis of pancreatitis and cancer through regulation of stromagenesis.

immunology↗

bioRxiv2021

Single-cell transcriptomics reveals a conserved metaplasia program in pancreatic injury

BACKGROUND & AIMSAcinar to ductal metaplasia (ADM) occurs in the pancreas in response to tissue injury and is a potential precursor for adenocarcinoma. The goal of these studies was to define the populations arising from ADM, the associated transcriptional changes, and markers of disease progression. METHODSAcinar cells were lineage-traced with enhanced yellow fluorescent protein (EYFP) to follow their fate upon injury. Transcripts of over 13,000 EYFP+ cells were determined using single-cell RNA sequencing (scRNA-seq). Developmental trajectories were generated. Data were compared to gastric metaplasia, KrasG12D-induced neoplasia, and human pancreatitis. Results were confirmed by immunostaining and electron microscopy. KrasG12D was expressed in injury-induced ADM using several inducible Cre drivers. Surgical specimens of chronic pancreatitis from 15 patients were evaluated by immunostaining. RESULTSscRNA-seq of ADM revealed emergence of a mucin/ductal population resembling gastric pyloric metaplasia. Lineage trajectories suggest that some pyloric metaplasia cells can generate tuft and enteroendocrine cells (EECs). Comparison to KrasG12D-induced ADM identifies populations associated with disease progression. Activation of KrasG12D expression in HNF1B+ or POU2F3+ ADM populations leads to neoplastic transformation and formation of MUC5AC+ gastric-pit-like cells. Human pancreatitis samples also harbor pyloric metaplasia with a similar transcriptional phenotype. CONCLUSIONSUnder conditions of chronic injury, acinar cells undergo a pyloric-type metaplasia to mucinous progenitor-like populations, which seed disparate tuft cell and EEC lineages. ADM-derived EEC subtypes are diverse. KrasG12D expression is sufficient to drive neoplasia from injury-induced ADM and offers an alternative origin for tumorigenesis. This program is conserved in human pancreatitis, providing insight into early events in pancreas diseases.

cell biology↗

bioRxiv2021

3'-Sulfated Lewis A is a Biomarker for Metaplastic and Oncogenic Transformation of Several Gastrointestinal Epithelia

IntroductionMultiple previous studies have shown the monoclonal antibody Das-1 (formerly called 7E12H12) specifically recognizes metaplastic and carcinomatous lesions in multiple organs of the gastrointestinal system (e.g. Barretts esophagus, intestinal-type metaplasia of the stomach, gastric adenocarcinoma, high-grade pancreatic intraepithelial neoplasm, and pancreatic ductal adenocarcinoma) as well as in other organs (bladder and lung carcinomas). Beyond being a useful biomarker in tissue, mAb Das-1 has recently proven to be more accurate than current paradigms for identifying cysts harboring advanced neoplasia. Though this antibody has been used extensively for clinical, basic science, and translational applications for decades, its epitope has remained elusive. MethodsIn this study, we chemically deglycosylated a standard source of antigen, which resulted in near complete loss of the signal as measured by western blot analysis. The epitope recognized by mAb Das-1 was determined by affinity to a comprehensive glycan array and validated by inhibition of a direct ELISA. ResultsThe epitope recognized by mAb Das-1 is 3-Sulfo-Lewis A (3-Sulfo-LeA). 3-Sulfo-LeA is broadly reexpressed across numerous GI epithelia and elsewhere only after metaplastic and carcinomatous transformation. Discussion3-Sulfo-LeA is a clinically important antigen that can be detected both intracellularly in tissue using immunohistochemistry and extracellularly in cyst fluid and serum by ELISA. The results open new avenues for tumorigenic risk stratification of various gastrointestinal lesions.

cancer biology↗