Biology subjects

Ross, S. R.

Publications and source records attributed to Ross, S. R..

13 recordsLinked to original sources

bioRxiv2026

SIRPA suppresses integrin-dependent virus endocytosis

New World arenaviruses (NWAs) that cause viral hemorrhagic fever, such as Junin virus, have few therapeutic options. Entry of these viruses into cells is mediated by binding to cell surface receptors, followed by endocytosis and trafficking to a low pH compartment. We showed previously that Signal Regulatory Protein Alpha (SIRPA), a critical cell surface receptor that inhibits macrophage phagocytic activity, decreases internalization by NWAs as well as other pathogenic RNA viruses that traffic to low pH compartments. Here we demonstrate that proteins involved in the SIRPA/integrin signaling axis, including Src homology region 2 (SH2)-containing protein tyrosine phosphatase 2 (SHP2), src family kinases (SFKs), particularly FYN, focal adhesion kinase (FAK), and alpha-integrin play a role in viral endocytosis and that SIRPA inhibits virus entry through blocking this pathway. In addition to defining a role for integrins in viral entry, these studies also provide additional insight into SIRPAs interference in processes dependent on integrin signaling, including phagocytosis. Moreover, using drugs that block the integrin signaling pathway in vitro and in vivo, we show that there are additional steps that may be targeted therapeutically for inhibiting infection by RNA viruses that traffic to acidic compartments.

cell biology↗

bioRxiv2026

IFI207 promotes antiviral responses by modulating STING ubiquitination and degradation

Aim2-like receptors (ALRs) play crucial roles in innate immune signaling pathways and demonstrate strong positive selection likely driven by pathogens. IFI207, an ALR found in all Mus species, enhances interaction with and stabilization of STING, contributing to the control of Murine Leukemia Virus (MLV) infection. We show here that IFI207 enhances the type 1 interferon response by inhibiting activation-induced K63-linked ubiquitination of STING, thereby preventing its recognition by hepatocyte growth factor-regulated tyrosine kinase substrate (HRS), a key component of the ESCRT complex, and its subsequent degradation in lysosomes. IFI207 promotes downstream signaling in the STING pathway in multiple cell types and moreover enhances the STING-dependent response to herpesvirus simplex 1 infection ex vivo and in vivo. We also show that IFI207 likely functions in dendritic cells to suppress MLV infection. Our study reveals that IFI207 acts as a modulator in the STING pathway, strengthening the hosts defense against viral infections and suggests that the expansion of the Alr locus in mice may have occurred in response to endemic viruses.

immunology↗

bioRxiv2024

DHX15 inhibits mouse APOBEC3 deamination

APOBEC3 family proteins are critical host factors that counteract and prevent the replication of retroviruses and other viruses through cytidine deamination. Human APOBEC3 proteins inactivate HIV-1 through the introduction of lethal mutations to viral genomes. In contrast, mouse APOBEC3 does not induce DNA hypermutation of murine retroviruses, although it retains functional cytidine deaminase activity. Why mouse APOBEC3 does not effectively deaminate murine retroviruses is still unknown. In this study, we found that the dead box helicase DHX15 interacts with mouse APOBEC3 and inhibits its deamination activity. DHX15 was packaged into murine leukemia virus (MLV) virions independent of its binding with APOBEC3. Moreover, DHX15 knockdown inhibited MLV replication and resulted in more G-to-A mutations in proviral DNA. Finally, DHX15 knockdown induced DNA damage in murine cells, suggesting that it plays a role in preserving genome integrity in cells expressing mouse APOBEC3 protein.

microbiology↗

bioRxiv2024

IFI207, a young and fast-evolving protein, controls retroviral replication via the STING pathway

Mammalian AIM-2-like receptor (ALR) proteins bind nucleic acids and initiate production of type I interferons or inflammasome assembly, thereby contributing to host innate immunity. In mice, the Alr locus is highly polymorphic at the sequence and copy number level and we show here, is one of the most dynamic regions of the genome. One rapidly evolving gene within this region, Ifi207, was introduced to the Mus genome by gene conversion or an unequal recombination event a few million years ago. Ifi207 has a large, distinctive repeat region that differs in sequence and length among Mus species and even closely related inbred Mus musculus strains. We show that IFI207 controls MLV infection in vivo and that it plays a role in the STING-mediated response to cGAMP, dsDNA, DMXXA and MLV. IFI207 binds to STING and inclusion of its repeat region appears to stabilize STING protein. The Alr locus and Ifi207 provide a clear example of the evolutionary innovation of gene function, possibly as a result of host-pathogen co-evolution. IMPORTANCEThe Red Queen hypothesis predicts that the arms race between pathogens and the host may accelerate evolution of both sides, and therefore cause higher diversity in virulence factors and immune-related proteins, respectively (1). The Alr gene family in mice has undergone rapid evolution in the last few million years and includes the creation of two novel members, MndaL and Ifi207. Ifi207 in particular became highly divergent, with significant genetic changes between highly related inbred mice. IFI207 protein acts in the STING pathway and contributes to anti-retroviral resistance via a novel mechanism. The data show that under the pressure of host-pathogen coevolution in a dynamic locus, gene conversion and recombination between gene family members creates new genes with novel and essential functions that play diverse roles in biological processes.

immunology↗

bioRxiv2023

Murine leukemia virus can infect non-dividing primary cells

Retroviral reverse transcription starts within the capsid and uncoating and reverse transcription are mutually dependent. There is still debate regarding the timing and cellular location of HIVs uncoating and reverse transcription and whether it occurs solely in the cytoplasm, nucleus or both. HIV can infect non-dividing cells because there is active transport of the preintegration complex (PIC) across the nuclear membrane, but Murine Leukemia Virus (MLV) is thought to depend on cell division for replication and whether MLV uncoating and reverse transcription is solely cytoplasmic has not been studied. Here, we used NIH3T3 and primary mouse dendritic cells to determine where the different stages of reverse transcription occur and whether cell division is needed for nuclear entry. Our data strongly suggest that in both NIH3T3 cells and dendritic cells (DCs), the initial step of reverse transcription occurs in the cytoplasm. However, we detected MLV RNA/DNA hybrid intermediates in the nucleus of dividing NIH3T3 cells and non-dividing DCs, suggesting that reverse transcription can continue after nuclear entry. We also found that the MLV PIC requires cell division to enter the nucleus of NIH3T3 cells. In contrast, we show that MLV can infect non-dividing primary DCs, although integration of MLV DNA in DCs still required the viral p12 protein. Knockdown of several nuclear pore proteins dramatically reduced the appearance of integrated MLV DNA in DCs but not NIH3T3 cells. Additionally, MLV capsid associates with the nuclear pore proteins NUP358 and NUP62 during infection. These findings suggest that simple retroviruses, like HIV, gain nuclear entry by traversing the nuclear pore complex in non-mitotic cells. Author SummaryIt is widely believed that gammaretroviruses like MLV require cell division to achieve nuclear entry and complete their replication. We show here that while this is true for rapidly dividing tissue culture cells, in quiescent cells like dendritic cells, the natural targets of MLV infection, the virus establishes infection without cell division. These studies show that the requirements for retrovirus infection depend on the cell type.

microbiology↗

bioRxiv2023

DNA damage primes hematopoietic stem cells for direct megakaryopoiesis

Hematopoietic stem cells (HSCs) reside in the bone marrow (BM), can self-renew, and generate all cells of the hematopoietic system.1 Most hematopoietic lineages arise through successive, increasingly lineage-committed progenitors. In contrast, megakaryocytes (MKs), hyperploid cells that generate platelets essential to hemostasis, can derive rapidly and directly from HSCs.2 The underlying mechanism is unknown however. Here we show that DNA damage and subsequent arrest in the G2 phase of the cell cycle rapidly induce MK commitment specifically in HSCs, but not in progenitors, through an initially predominantly post-transcriptional mechanism. Cycling HSCs show extensive replication-induced DNA damage associated with uracil misincorporation in vivo and in vitro. Consistent with this notion, thymidine attenuated DNA damage, rescued HSC maintenance and reduced the generation of CD41+ MK-committed HSCs in vitro. Similarly, overexpression of the dUTP-scavenging enzyme, dUTPase, enhanced in vitro maintenance of HSCs. We conclude that a DNA damage response drives direct megakaryopoiesis and that replication stress-induced direct megakaryopoiesis, at least in part caused by uracil misincorporation, is a barrier to HSC maintenance in vitro. DNA damage-induced direct megakaryopoiesis may allow rapid generation of a lineage essential to immediate organismal survival, while simultaneously removing damaged HSCs and potentially avoiding malignant transformation of self-renewing stem cells.

cell biology↗

bioRxiv2023

IFI207, a young and fast-evolving antiviral factor, stabilizes STING

Mammalian ALR proteins bind nucleic acids and initiate production of type I interferons or inflammasome assembly, thereby contributing to host innate immunity. ALRs are encoded at a single genetic locus. In mice, the Alr locus is highly polymorphic at the sequence and copy number level. We suggest that one rapidly evolving member of the Alr family, Ifi207, was introduced to the Mus genome by a recent recombination event. Ifi207 has a large, distinctive repeat region that differs in sequence and length in different Mus strains. We show that IFI207 plays a key role in the STING-mediated response to cGAMP, DNA, and MLV, and that IFI207 controls MLV infection in vivo. Uniquely, IFI207 acts by stabilizing STING protein via its repeat region. Our studies suggest that under the pressure of host-pathogen coevolution, in a dynamic locus such as the Alr, recombination between gene family members creates new genes with novel and essential functions that play diverse roles in biological processes.

microbiology↗

bioRxiv2022

The cytidine deaminase APOBEC3G drives cancer mutagenesis and clonal evolution in bladder cancer

Mutagenic processes leave distinct signatures in cancer genomes. The mutational signatures attributed to APOBEC3 cytidine deaminases are pervasive in human cancers. However, data linking individual APOBEC3 proteins to cancer mutagenesis in vivo are limited. Here, we show that transgenic expression of human APOBEC3G promotes mutagenesis, genomic instability, and kataegis, leading to shorter survival in a murine bladder cancer model. Acting as mutagenic fuel, APOBEC3G increases the clonal diversity of bladder cancers, driving divergent cancer evolution. We characterize the single base substitution signature induced by APOBEC3G in vivo, showing the induction of a mutational signature different from that caused by APOBEC3A and APOBEC3B. Analysis of thousands of human cancers reveals the contribution of APOBEC3G to the mutational profiles of multiple cancer types, including bladder cancer. Our findings define the role of APOBEC3G in cancer mutagenesis and clonal heterogeneity. These results potentially inform future therapeutic efforts that restrict tumor evolution.

genomics↗

bioRxiv2021

Repair of APOBEC3G-mutated retroviral DNA in vivo is facilitated by the host enzyme uracil DNA glycosylase 2

Apolipoprotein B mRNA Editing Enzyme Catalytic Subunit 3 (APOBEC3) proteins are critical for the control of infection by retroviruses. These proteins deaminate cytidines in negative strand DNA during reverse transcription, leading to G to A changes in coding strands. Uracil DNA glycosylase (UNG) is a host enzyme that excises uracils in genomic DNA, which the base excision repair machinery then repairs. Whether UNG removes uracils found in retroviral DNA after APOBEC3-mediated mutation is not clear, and whether this occurs in vivo has not been demonstrated. To determine if UNG plays a role in the repair of retroviral DNA, we used APOBEC3G (A3G) transgenic mice which we showed previously had extensive deamination of murine leukemia virus (MLV) proviruses. The A3G transgene was crossed onto an UNG and mouse APOBEC3 knockout background (UNG-/-APO-/-) and the mice were infected with MLV. We found that virus infection levels were decreased in A3G UNG-/-APO-/- compared to A3G APO-/- mice. Deep sequencing of the proviruses showed that there were significantly higher levels of G-to-A mutations in proviral DNA from A3G transgenic UNG-/-APO-/- than A3G transgenic APO-/- mice, suggesting that UNG plays a role in the repair of uracil-containing proviruses. In in vitro studies, we found that cytoplasmic viral DNA deaminated by APOBEC3G was uracilated. In the absence of UNG, the uracil-containing proviruses integrated at higher levels into the genome than did those made in the presence of UNG. Thus, UNG also functions in the nucleus prior to integration by nicking uracil-containing viral DNA, thereby blocking integration. These data show that UNG plays a critical role in the repair of the damage inflicted by APOBEC3 deamination of reverse-transcribed DNA. ImportanceWhile APOBEC3-mediated mutation of retroviruses is well-established, what role the host base excision repair enzymes play in correcting these mutations is not clear. This question is especially difficult to address in vivo. Here, we use a transgenic mouse developed by our lab that expresses human APOBEC3G and also lacks the endogenous uracil DNA glycosylase (Ung) gene, and show that UNG removes uracils introduced by this cytidine deaminase in MLV reverse transcripts, thereby reducing G-to-A mutations in proviruses. Furthermore, our data suggest that UNG removes uracils at two stages in infection - in unintegrated nuclear viral reverse transcribed DNA, resulting in its degradation and second, in integrated proviruses, resulting in their repair. These data suggest that retroviruses damaged by host cytidine deaminases take advantage of the host DNA repair system to overcome this damage.

microbiology↗

bioRxiv2021

DDX41 is needed for pre-and post-natal hematopoietic stem cell differentiation in mice

DDX41 is a tumor suppressor frequently mutated in human myeloid neoplasms. DDX41 binds to DNA/RNA hybrids and interacts with spliceosome component (1, 2). How it affects hematopoiesis is still unclear. Using a knockout mouse model, we demonstrate that DDX41 is required for mouse hematopoietic stem and progenitor cell (HSPC) survival and differentiation. Lack of DDX41 particularly affected myeloid progenitor development, starting at embryonic day 13.5. DDX41-deficient fetal liver and adult bone marrow (BM) cells were unable to rescue mice from lethal irradiation after transplantation. DDX41 knockout stem cells were also defective in ex vivo colony forming assays. RNASeq analysis of lineage-negative, cKit+Sca1+ cells isolated from fetal liver demonstrated that the expression of many genes associated with hematopoietic differentiation were altered in DDX41 knockout cells. Furthermore, altered splicing of genes involved in key biological processes were observed. Our data reveal a critical role for DDX41 in HSPC differentiation and myeloid progenitor development, likely through its regulation of gene expression programs and splicing. SignificanceDDX41 is a tumor suppressor in hematologic malignancies. However, whether DDX41 functions in hematopoiesis and myeloid cell differentiation is not known. Here we show that in mice, loss of DDX41 in hematopoietic stem cells (HSCs) leads to defects in hematopoietic development. The myeloid lineage was particularly affected as early as pre-natal stages. Transcriptional profiling of embryonic HSCs revealed that there were global changes in gene expression and splicing due to lack of DDX41. Collectively, the study uncovers a new function of DDX41 in HSC differentiation and could provide molecular targets for treatment of myeloid differentiation disorders.

developmental biology↗

bioRxiv2019

APOBEC3A Catalyzes Mutation and Drives Carcinogenesis In Vivo

The APOBEC3 family of antiviral DNA cytosine deaminases is implicated as the second largest source of mutation in cancer. This mutational process may be a causal driver or inconsequential passenger to the overall tumor phenotype. We show that human APOBEC3A expression in murine colon and liver tissues increases tumorigenesis. All other APOBEC3 family members including APOBEC3B did not as strongly promote liver tumor formation. DNA sequences from APOBEC3A-expressing animals display hallmark APOBEC signature mutations in TCA/T motifs. Bioinformatic comparisons of the observed APOBEC3A mutation signature in murine tumors, previously reported APOBEC3A and APOBEC3B mutation signatures in yeast, and reanalyzed APOBEC mutation signatures in human tumor data sets support cause-and-effect relationships for APOBEC3A-catalyzed deamination and mutagenesis in driving multiple human cancers.

cancer biology↗

bioRxiv2018

Human APOBEC3G prevents emergence of infectious endogenous retrovirus in mice

Endogenous retroviruses (ERV) are found throughout vertebrate genomes and failure to silence their activation can have deleterious consequences on the host. Introduction of mutations that subsequently prevent transcription of ERV loci is therefore an indispensable cell-intrinsic defense mechanism that maintains the integrity of the host genome. Abundant in vitro and in silico evidence have revealed that APOBEC3 cytidine-deaminases, including human APOBEC3G (hA3G) can potently restrict retrotransposition; yet in vivo data demonstrating such activity is lacking, particularly since no replication competent human ERV has been identified. In mice deficient for Toll-like receptor 7 (TLR7), transcribed ERV loci can recombine and generate infectious ERV. In this study, we show that mice deficient in the only copy of Apobec3 in the genome did not have spontaneous reactivation of ERVs, nor elevated ERV reactivation when crossed to Tlr7-/- mice. In contrast, expression of a human APOBEC3G transgene abrogated emergence of infectious ERV in the Tlr7-/- background. No ERV RNA was detected in the plasma of hA3G+Apobec3-/-Tlr7-/- mice, and infectious ERV virions could not be amplified through co-culture with permissive cells. These data reveal that hA3G can potently restrict active ERV in vivo, and suggest that the expansion of the APOBEC3 locus in primates has helped restrict ERV reactivation in the human genome.\n\nImportanceAlthough APOBEC3 proteins are known to be important antiviral restriction factors in both mice and humans, their roles in the restriction of endogenous retroviruses (ERV) have been limited to in vitro studies. Here, we report that human APOBEC3G expressed as a transgene in mice prevents the emergence of infectious ERV from endogenous loci. This study reveals that APOBEC3G can powerfully restrict active retrotransposons in vivo and demonstrates how ectopic expression of human factors in transgenic mouse models can be used to investigate host mechanisms that inhibit retrotransposons and reinforce genomic integrity.

immunology↗

bioRxiv2018

DDX41 recognizes RNA/DNA retroviral reverse transcripts and is critical for in vivo control of MLV infection

Host recognition of viral nucleic acids generated during infection leads to the activation of innate immune responses essential for early control of virus. Retrovirus reverse transcription creates numerous potential ligands for cytosolic host sensors that recognize foreign nucleic acids, including single-stranded RNA (ssRNA), RNA/DNA hybrids and double stranded DNA (dsDNA). We and others recently showed that the sensors cyclic GMP-AMP synthase (cGAS), dead-box helicase 41 (DDX41) and members of the Aim2-like receptor (ALR) family participate in the recognition of retroviral reverse transcripts. However, why multiple sensors might be required and their relative importance in in vivo control of retroviral infection is not known. Here we show that DDX41 primarily senses the DNA/RNA hybrid generated at the first step of reverse transcription, while cGAS recognizes dsDNA generated at the next step. We also show that both DDX41 and cGAS are needed for the anti-retroviral innate immune response to MLV and HIV in primary mouse macrophages and dendritic cells (DC). Using mice with macrophage- or -specific knockout of the DDX41 gene, we show that DDX41 sensing in DCs but not macrophages was critical for controlling in vivo MLV infection. This suggests that DCs are essential in vivo targets for infection, as well as for initiating the antiviral response. Our work demonstrates that the innate immune response to retrovirus infection depends on multiple host nucleic acid sensors that recognize different reverse transcription intermediates.\n\nImportanceViruses are detected by many different host sensors of nucleic acid, which in turn trigger innate immune responses, such as type I IFN production, required to control infection. We show here that at least two sensors are needed to initiate a highly effective innate immune response to retroviruses - DDX41, which preferentially senses the RNA/DNA hybrid generated at the first step of retrovirus replication and cGAS, which recognizes double-stranded DNA generated at the 2nd step. Importantly, we demonstrate using mice lacking DDX41 or cGAS, that both sensors are needed for the full antiviral response needed to control in vivo MLV infection. These findings underscore the need for multiple host factors to counteract retroviral infection.

microbiology↗