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1.
Elife ; 122023 11 07.
Article in English | MEDLINE | ID: mdl-37934570

ABSTRACT

ZMYM2 is a zinc finger transcriptional regulator that plays a key role in promoting and maintaining cell identity. It has been implicated in several diseases such as congenital anomalies of the kidney where its activity is diminished and cancer where it participates in oncogenic fusion protein events. ZMYM2 is thought to function through promoting transcriptional repression and here we provide more evidence to support this designation. Here we studied ZMYM2 function in human cells and demonstrate that ZMYM2 is part of distinct chromatin-bound complexes including the established LSD1-CoREST-HDAC1 corepressor complex. We also identify new functional and physical interactions with ADNP and TRIM28/KAP1. The ZMYM2-TRIM28 complex forms in a SUMO-dependent manner and is associated with repressive chromatin. ZMYM2 and TRIM28 show strong functional similarity and co-regulate a large number of genes. However, there are no strong links between ZMYM2-TRIM28 binding events and nearby individual gene regulation. Instead, ZMYM2-TRIM28 appears to regulate genes in a more regionally defined manner within TADs where it can directly regulate co-associated retrotransposon expression. We find that different types of ZMYM2 binding complex associate with and regulate distinct subclasses of retrotransposons, with ZMYM2-ADNP complexes at SINEs and ZMYM2-TRIM28 complexes at LTR elements. We propose a model whereby ZMYM2 acts directly through retrotransposon regulation, which may then potentially affect the local chromatin environment and associated coding gene expression.


Subject(s)
DNA Transposable Elements , Retroelements , Humans , Zinc Fingers , Chromatin , Co-Repressor Proteins , DNA-Binding Proteins , Transcription Factors
2.
Am J Hum Genet ; 107(4): 727-742, 2020 10 01.
Article in English | MEDLINE | ID: mdl-32891193

ABSTRACT

Congenital anomalies of the kidney and urinary tract (CAKUT) constitute one of the most frequent birth defects and represent the most common cause of chronic kidney disease in the first three decades of life. Despite the discovery of dozens of monogenic causes of CAKUT, most pathogenic pathways remain elusive. We performed whole-exome sequencing (WES) in 551 individuals with CAKUT and identified a heterozygous de novo stop-gain variant in ZMYM2 in two different families with CAKUT. Through collaboration, we identified in total 14 different heterozygous loss-of-function mutations in ZMYM2 in 15 unrelated families. Most mutations occurred de novo, indicating possible interference with reproductive function. Human disease features are replicated in X. tropicalis larvae with morpholino knockdowns, in which expression of truncated ZMYM2 proteins, based on individual mutations, failed to rescue renal and craniofacial defects. Moreover, heterozygous Zmym2-deficient mice recapitulated features of CAKUT with high penetrance. The ZMYM2 protein is a component of a transcriptional corepressor complex recently linked to the silencing of developmentally regulated endogenous retrovirus elements. Using protein-protein interaction assays, we show that ZMYM2 interacts with additional epigenetic silencing complexes, as well as confirming that it binds to FOXP1, a transcription factor that has also been linked to CAKUT. In summary, our findings establish that loss-of-function mutations of ZMYM2, and potentially that of other proteins in its interactome, as causes of human CAKUT, offering new routes for studying the pathogenesis of the disorder.


Subject(s)
DNA-Binding Proteins/genetics , Epigenesis, Genetic , Forkhead Transcription Factors/genetics , Mutation , Repressor Proteins/genetics , Transcription Factors/genetics , Urinary Tract/metabolism , Urogenital Abnormalities/genetics , Amphibian Proteins/antagonists & inhibitors , Amphibian Proteins/genetics , Amphibian Proteins/metabolism , Animals , Case-Control Studies , Child , Child, Preschool , DNA-Binding Proteins/metabolism , Family , Female , Forkhead Transcription Factors/metabolism , Heterozygote , Humans , Infant , Larva/genetics , Larva/growth & development , Larva/metabolism , Male , Mice , Mice, Knockout , Morpholinos/genetics , Morpholinos/metabolism , Pedigree , Protein Binding , Repressor Proteins/metabolism , Transcription Factors/metabolism , Urinary Tract/abnormalities , Urogenital Abnormalities/metabolism , Urogenital Abnormalities/pathology , Exome Sequencing , Xenopus
3.
Elife ; 92020 05 11.
Article in English | MEDLINE | ID: mdl-32391791

ABSTRACT

Herpesviruses acquire their membrane envelopes in the cytoplasm of infected cells via a molecular mechanism that remains unclear. Herpes simplex virus (HSV)-1 proteins pUL7 and pUL51 form a complex required for efficient virus envelopment. We show that interaction between homologues of pUL7 and pUL51 is conserved across human herpesviruses, as is their association with trans-Golgi membranes. We characterized the HSV-1 pUL7:pUL51 complex by solution scattering and chemical crosslinking, revealing a 1:2 complex that can form higher-order oligomers in solution, and we solved the crystal structure of the core pUL7:pUL51 heterodimer. While pUL7 adopts a previously-unseen compact fold, the helix-turn-helix conformation of pUL51 resembles the cellular endosomal complex required for transport (ESCRT)-III component CHMP4B and pUL51 forms ESCRT-III-like filaments, suggesting a direct role for pUL51 in promoting membrane scission during virus assembly. Our results provide a structural framework for understanding the role of the conserved pUL7:pUL51 complex in herpesvirus assembly.


Most people suffer from occasional cold sores, which are caused by the herpes simplex virus. This virus causes infections that last your entire life, but for the most part it lies dormant in your cells and reactivates only at times of stress. When it reactivates, the virus manipulates host cells to make new virus particles that may spread the infection to other people. Like many other viruses, herpes simplex viruses also steal jelly-like structures known as membranes from their host cells to form protective coats around new virus particles. In cells from humans and other animals, proteins belonging to a molecular machine known as ESCRT form filaments that bend and break membranes as the cells require. Many viruses hijack the ESCRT machinery to wrap membranes around new virus particles. However, herpes simplex viruses do not follow the usual rules for activating this machine. Instead, they rely on two viral proteins called pUL7 and pUL51 to hot-wire the ESCRT machinery. Previous studies have shown that these two proteins bind to each other, but it remained unclear how they work. Butt et al. used a combination of biochemical and biophysical techniques to solve the three-dimensional structures of pUL7 and pUL51 when bound to each other. The experiments determined that the structure of pUL51 resembles the structures of different components in the ESCRT machinery. Like the ESCRT proteins, pUL51 formed filaments, suggesting that pUL51 bends membranes in cells and that pUL7 blocks it from doing so until the time is right. Further experiments showed that the equivalents of pUL7 and pUL51 in other members of the herpes virus family also bind to each other in a similar way. These findings reveal that herpes simplex viruses and their close relatives have evolved a different strategy than many other viruses to steal membranes from host cells. Interfering with this mechanism may provide new avenues for designing drugs or improving vaccines against these viruses. The pUL7 and pUL51 proteins may also inspire new tools in biotechnology that could precisely control the shapes of biological membranes.


Subject(s)
Herpesvirus 1, Human/physiology , Phosphoproteins/chemistry , Phosphoproteins/genetics , Viral Matrix Proteins/chemistry , Viral Matrix Proteins/genetics , Viral Proteins/chemistry , Viral Proteins/genetics , Virus Assembly , HEK293 Cells , HeLa Cells , Herpes Simplex/virology , Herpesvirus 1, Human/chemistry , Humans , Models, Molecular , Protein Binding , Protein Interaction Domains and Motifs , Protein Structure, Quaternary , Protein Structure, Tertiary , Viral Proteins/metabolism , Virus Replication , trans-Golgi Network
4.
J Virol ; 91(2)2017 Jan 15.
Article in English | MEDLINE | ID: mdl-27852850

ABSTRACT

The tegument of herpesviruses is a highly complex structural layer between the nucleocapsid and the envelope of virions. Tegument proteins play both structural and regulatory functions during replication and spread, but the interactions and functions of many of these proteins are poorly understood. Here we focus on two tegument proteins from herpes simplex virus 1 (HSV-1), pUL7 and pUL51, which have homologues in all other herpesviruses. We have now identified that HSV-1 pUL7 and pUL51 form a stable and direct protein-protein interaction, their expression levels rely on the presence of each other, and they function as a complex in infected cells. We demonstrate that expression of the pUL7-pUL51 complex is important for efficient HSV-1 assembly and plaque formation. Furthermore, we also discovered that the pUL7-pUL51 complex localizes to focal adhesions at the plasma membrane in both infected cells and in the absence of other viral proteins. The expression of pUL7-pUL51 is important to stabilize focal adhesions and maintain cell morphology in infected cells and cells infected with viruses lacking pUL7 and/or pUL51 round up more rapidly than cells infected with wild-type HSV-1. Our data suggest that, in addition to the previously reported functions in virus assembly and spread for pUL51, the pUL7-pUL51 complex is important for maintaining the attachment of infected cells to their surroundings through modulating the activity of focal adhesion complexes. IMPORTANCE: Herpesviridae is a large family of highly successful human and animal pathogens. Virions of these viruses are composed of many different proteins, most of which are contained within the tegument, a complex structural layer between the nucleocapsid and the envelope within virus particles. Tegument proteins have important roles in assembling virus particles as well as modifying host cells to promote virus replication and spread. However, little is known about the function of many tegument proteins during virus replication. Our study focuses on two tegument proteins from herpes simplex virus 1 that are conserved in all herpesviruses: pUL7 and pUL51. We demonstrate that these proteins directly interact and form a functional complex that is important for both virus assembly and modulation of host cell morphology. Further, we identify for the first time that these conserved herpesvirus tegument proteins localize to focal adhesions in addition to cytoplasmic juxtanuclear membranes within infected cells.


Subject(s)
DNA Helicases/metabolism , DNA Primase/metabolism , Herpes Simplex/metabolism , Herpes Simplex/virology , Herpesvirus 1, Human/physiology , Multiprotein Complexes/metabolism , Viral Matrix Proteins/metabolism , Viral Proteins/metabolism , Animals , Chlorocebus aethiops , DNA Helicases/genetics , DNA Primase/genetics , Gene Expression Regulation, Viral , HEK293 Cells , Herpesvirus 1, Human/ultrastructure , Humans , Protein Binding , Protein Transport , Vero Cells , Viral Matrix Proteins/genetics , Viral Proteins/genetics , Virus Assembly
5.
Viruses ; 7(9): 5084-114, 2015 Sep 18.
Article in English | MEDLINE | ID: mdl-26393641

ABSTRACT

Alphaherpesviruses like herpes simplex virus are large DNA viruses characterized by their ability to establish lifelong latent infection in neurons. As for all herpesviruses, alphaherpesvirus virions contain a protein-rich layer called "tegument" that links the DNA-containing capsid to the glycoprotein-studded membrane envelope. Tegument proteins mediate a diverse range of functions during the virus lifecycle, including modulation of the host-cell environment immediately after entry, transport of virus capsids to the nucleus during infection, and wrapping of cytoplasmic capsids with membranes (secondary envelopment) during virion assembly. Eleven tegument proteins that are conserved across alphaherpesviruses have been implicated in the formation of the tegument layer or in secondary envelopment. Tegument is assembled via a dense network of interactions between tegument proteins, with the redundancy of these interactions making it challenging to determine the precise function of any specific tegument protein. However, recent studies have made great headway in defining the interactions between tegument proteins, conserved across alphaherpesviruses, which facilitate tegument assembly and secondary envelopment. We summarize these recent advances and review what remains to be learned about the molecular interactions required to assemble mature alphaherpesvirus virions following the release of capsids from infected cell nuclei.


Subject(s)
Alphaherpesvirinae/physiology , Virus Assembly , Models, Biological , Protein Binding , Viral Structural Proteins/metabolism
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