Gregory Fonseca

McGill University | McGill · Department of Medicine

Regulation of gene expression requires the combinatorial binding of sequence-specific transcription factors (TFs) at promoters and enhancers. Prior studies showed that alterations in the spacing between TF binding sites can influence promoter and enhancer activity. However, the relative importance of TF spacing alterations resulting from naturally...

Viruses alter a multitude of host-cell processes to create a more optimal environment for viral replication. This includes altering metabolism to provide adequate substrates and energy required for replication. Typically, viral infections induce a metabolic phenotype resembling the Warburg effect, with an upregulation of glycolysis and a concurrent...

Regulation of gene expression requires the combinatorial binding of sequence-specific transcription factors (TFs) at promoters and enhancers. Single nucleotide polymorphisms (SNPs) and short insertions and deletions (InDels) can influence gene expression by altering the sequences of TF binding sites. Prior studies also showed that alterations in th...

The lung is inhabited by resident alveolar and interstitial macrophages as well as monocytic cells that survey lung tissues. Each cell type plays distinct functional roles under homeostatic and inflammatory conditions, but mechanisms establishing their molecular identities and functional potential remain poorly understood. In the present study, sys...

Cerebral cavernous malformations (CCMs) are dilated capillaries causing epilepsy and stroke. Inheritance of a heterozygous mutation in CCM3/PDCD10 is responsible for the most aggressive familial form of the disease. Here we studied the differences and commonalities between the transcriptomes of microdissected lesional neurovascular units (NVUs) fro...

The purpose of this study was to determine important genes, functions, and networks contributing to the pathobiology of cerebral cavernous malformation (CCM) from transcriptomic analyses across 3 species and 2 disease genotypes. Sequencing of RNA from laser microdissected neurovascular units of 5 human surgically resected CCM lesions, mouse brain m...

Mechanisms by which members of the AP-1 family of transcription factors play non-redundant biological roles despite recognizing the same DNA sequence remain poorly understood. To address this question, here we investigate the molecular functions and genome-wide DNA binding patterns of AP-1 family members in primary and immortalized mouse macrophage...

Mechanisms by which members of the AP-1 family of transcription factors play both redundant and non-redundant biological roles despite recognizing the same DNA sequence remain poorly understood. To address this question, we investigated the molecular functions and genome-wide DNA binding patterns of AP-1 family members in macrophages. ChIP-sequenci...

Non-coding genetic variation is a major driver of phenotypic diversity and allows the investigation of mechanisms that control gene expression. Here, we systematically investigated the effects of >50 million variations from five strains of mice on mRNA, nascent transcription, transcription start sites, and transcription factor binding in resting an...

KRIT1 mutations are the most common cause of cerebral cavernous malformation (CCM). AcuteKrit1gene inactivation in mouse brain microvascular endothelial cells (BMECs) changes expression of multiple genes involved in vascular development. These changes include suppression ofThbs1, which encodes thrombospondin1 (TSP1) and has been ascribed to KLF2- a...

Genome-Wide Approaches to Defining Macrophage Identity and Function, Page 1 of 2 Abstract Macrophages play essential roles in the response to injury and infection and contribute to the development and/or homeostasis of the various tissues they reside in. Conversely, macrophages also influence the pathogenesis of metabolic, neurodegenerative, and...

The oncoproteins of the small DNA tumor viruses interact with a plethora of cellular regulators to commandeer control of the infected cell. During infection, adenovirus E1A deregulates cAMP signalling and repurposes it for activation of viral gene expression. We show that E1A structurally and functionally mimics a cellular A-kinase anchoring protei...

Related to Fig 6: Co-staining of PKA and viral replication centres during HAdV-5 infection. A549 cells were infected with the indicated virus (MOI 5) and were subsequently fixed, permeabilized and stained with antibodies specific for the indicated PKA subunits or HAdV-5 DNA-binding protein (DBP) and DAPI as indicated. Images were acquired on a Niko...

Related to Fig 7: PKA is recruited to HAdV early gene promoters in an E1A-dependent manner. A549 cells were infected with the indicated viruses at an MOI of 5 and harvested 20 hours post-infection. Chromatin immunoprecipitation (ChIP) was performed with antibodies specific for the indicated proteins. DNA was probed via qPCR for the presence of mult...

List of silencing RNAs used in this study. (DOCX)

List of antibodies used in this study. (DOCX)

List of primers used in this study. (DOCX)

Related to Fig 6: Subcellular localization of RIIα and Cα during HAdV-5 infection and localization of all PKA subunits in HEK293 cells. (A) A549 cells were infected with either WT HAdV-5 (dl309), ΔE1A virus (dl312) or a virus lacking PKA-binding (dl1101; Δ4–25). Cells were fixed, permeabilized and stained for confocal immunofluorescence. RIIα appea...

Related to Fig 6: HAdV-4 E1A does not relocalize PKA subunits. A549 cells were transfected with EGFP-tagged constructs for full-length HAdV-5 or HAdV-4 E1A. Cells were fixed, permeabilized and stained with antibodies for PKA subunits and DAPI as indicated. Unlike HAdV-5 E1A, HAdV-4 E1A was unable to noticeably relocalize PKA, suggesting that an AKA...

Related to Fig 7: PKA is required for WT levels of HAdV-5 protein production. A549 cells were treated with control siRNA or siRNA specific for PKA subunits and infected with WT HAdV-5 (dl309; MOI of 5). Cells were harvested at 12, 24, and 36 hr post-infection and viral protein production was assayed by western blot using antibodies against represen...

Related to Fig 3: HAdV-4 E1A is not predicted to form a helix that is capable of binding PKA in an equivalent manner as the HAdV-5 E1A AKAP-like sequence. (A) The PSI-PRED protein sequence analysis workbench was used to predict the helical propensity of the N-terminal regions of HAdV-5 and HAdV-4 E1A. Although both sequences are predicted to form h...

Macrophages reside in essentially all tissues of the body and play key roles in innate and adaptive immune responses. Distinct populations of tissue macrophages also acquire context-specific functions that are important for normal tissue homeostasis. To investigate mechanisms responsible for tissue-specific functions, we analyzed the transcriptomes...

During infection by human adenovirus (HAdV), the proteins encoded by the early region 1A (E1A) gene bind and appropriate components of the cellular transcriptional machinery to activate viral early genes transcription. Previously, we identified roles for the hBre1 and the hPaf1 complexes in E1A mediated transcriptional activation of HAdV early gene...

The early region 1A (E1A) of human adenovirus types 2 and 5 are differentially spliced to yield five distinct mRNAs that encode different proteins. The smallest E1A RNA transcript encodes a 55 residue (R) protein that shares only 28 amino acid residues in common with the other E1A proteins. Even though it is the most abundant E1A transcript at late...

Human Adenovirus E1A makes extensive connections with the cellular protein interaction network. By doing so, E1A can manipulate many cellular programs including cell cycle progression. Through these reprogramming events, E1A functions as a growth promoting oncogene and has been used extensively to investigate mechanisms contributing to oncogenesis....

Supplementary materials including Figure S1, S2, S3, S4 and Table S1 & S2 (PDF)

Upon infection, human adenovirus (HAdV) must activate the expression of its early genes to reprogram the cellular environment to support virus replication. This activation is orchestrated in large part by the first HAdV gene expressed during infection, early region 1A (E1A). E1A binds and appropriates components of the cellular transcriptional mach...

Overcoming the cellular type I interferon (IFN) host defense response is critical for a virus to ensure successful infection. Investigating the effects of human adenovirus (HAdV) infection on global cellular histone posttranslational modification (hPTM), we discovered that virus infection-induced activation of IFN signaling triggers a global increa...

The largest isoform of adenovirus early region 1A (E1A) contains a unique region termed conserved region 3 (CR3). This region activates viral gene expression by recruiting cellular transcription machinery to the early viral promoters. Recent studies have suggested that there is an optimal level of E1A-dependent transactivation required by human ade...

Early region 1A (E1A) of human adenovirus (HAdV) has been the focus of over 30 years of investigation and is required for the oncogenic capacity of HAdV in rodents. Alternative splicing of the E1A transcript generates mRNAs encoding multiple E1A proteins. The 55-residue (55R) E1A protein, which is encoded by the 9S mRNA, is particularly interesting...

The adenovirus E1A proteins function via protein-protein interactions. By making many connections with the cellular protein network, individual modules of this virally encoded hub reprogram numerous aspects of cell function and behavior. Although many of these interactions have been thoroughly studied, those mediated by the C-terminal region of E1A...

Hub proteins have central roles in regulating cellular processes. By targeting a single cellular hub, a viral oncogene may gain control over an entire module in the cellular interaction network that is potentially comprised of hundreds of proteins. The adenovirus E1A oncoprotein is a viral hub that interacts with many cellular hub proteins by short...

C-terminal binding protein (CtBP) binds to adenovirus early region 1A (AdE1A) through a highly conserved PXDLS motif close to the C terminus. We now have demonstrated that CtBP1 also interacts directly with the transcriptional activation domain (conserved region 3 [CR3]) of adenovirus type 5 E1A (Ad5E1A) and requires the integrity of the entire CR3...

Viruses are obligate intracellular parasites. Their genomes are not large enough to encode all the functions required to independently produce progeny; hence, viruses are absolutely dependent on host cell functions. Mechanistically, these host cell processes in eukaryotes are founded on an exquisitely complex series of molecular interactions. In pa...