Ananda Roy - Academia.edu (original) (raw)

Papers by Ananda Roy

Frontiers in Immunology

Transcriptional regulation of cells in the immune system must be strictly controlled at multiple ... more Transcriptional regulation of cells in the immune system must be strictly controlled at multiple levels to ensure that a proper immune response is elicited only when required. Analysis in bulk, or ensemble of cells, provides a wealth of important information leading to a better understanding of the various molecular steps and mechanisms involved in regulating gene expression in immune cells. However, given the substantial heterogeneity of these cells, it is imperative now to decipher these mechanisms at a single cell level. Here I bring together several recent examples to review our understanding of transcriptional regulation of the immune system via single cell analysis and to further illustrate the immense power of such analyses to interrogate immune cell heterogeneity.

Frontiers in Immunology, Jun 14, 2019

Transcriptional regulation of cells in the immune system must be strictly controlled at multiple ... more Transcriptional regulation of cells in the immune system must be strictly controlled at multiple levels to ensure that a proper immune response is elicited only when required. Analysis in bulk, or ensemble of cells, provides a wealth of important information leading to a better understanding of the various molecular steps and mechanisms involved in regulating gene expression in immune cells. However, given the substantial heterogeneity of these cells, it is imperative now to decipher these mechanisms at a single cell level. Here I bring together several recent examples to review our understanding of transcriptional regulation of the immune system via single cell analysis and to further illustrate the immense power of such analyses to interrogate immune cell heterogeneity.

Molecular Biology of the Cell

The adult human body is composed of nearly 37 trillion cells, each with potentially unique molecu... more The adult human body is composed of nearly 37 trillion cells, each with potentially unique molecular characteristics. This Perspective describes some of the challenges and opportunities faced in mapping the molecular characteristics of these cells in specific regions of the body and highlights areas for international collaboration toward the broader goal of comprehensively mapping the human body with cellular resolution.

Science Advances

The NIH Roadmap Epigenomics Program was launched to deliver reference epigenomic data from human ... more The NIH Roadmap Epigenomics Program was launched to deliver reference epigenomic data from human tissues and cells, develop tools and methods for analyzing the epigenome, discover novel epigenetic marks, develop methods to manipulate the epigenome, and determine epigenetic contributions to diverse human diseases. Here, we comment on the outcomes from this program: the scientific contributions made possible by a consortium approach and the challenges, benefits, and lessons learned from this group science effort.

Science advances, 2018

It has become exceedingly important to understand the precise molecular profiles of the nearly 40... more It has become exceedingly important to understand the precise molecular profiles of the nearly 40 trillion cells in an adult human because of their role in determining health, disease, and therapeutic outcome. The National Institutes of Health (NIH) Common Fund-supported Single Cell Analysis Program (SCAP) was designed to address this challenge. In this review, we outline the original program goals and provide a perspective on the impact of the program as a catalyst for exploration of heterogeneity of human tissues at the cellular level. We believe that the technological advances in single-cell RNA sequencing and multiplexed imaging combined with computational methods made by this program will undoubtedly have an impact on broad and robust applications of single-cell analyses in both health and disease research.

Trends in molecular medicine, 2017

The biochemical properties of the signal-induced multifunctional transcription factor II-I (TFII-... more The biochemical properties of the signal-induced multifunctional transcription factor II-I (TFII-I) indicate that it is involved in a variety of gene regulatory processes. Although gene ablation in murine models and cell-based assays show that it is encoded by an essential gene, GTF2I/Gtf2i, its physiologic role in human disorders was relatively unknown until recently. Novel studies show that it is involved in an array of human diseases including neurocognitive disorders, systemic lupus erythematosus (SLE), and cancer. Here I bring together these diverse observations to illustrate its multiple pathophysiologic functions and further conjecture on how these could be related to its known biochemical properties. I expect that a better understanding of these 'structure-function' relationships would lead to future diagnostic and/or therapeutic potential.

Molecular and Cellular Biology, Sep 1, 1998

In our effort to understand the transcriptional regulation of naturally occurring TATA-less but i... more In our effort to understand the transcriptional regulation of naturally occurring TATA-less but initiator (Inr)-containing genes, we have employed the murine T-cell receptor V␤ 5.2 promoter as a model. Here we show by transient-transfection assays that the Inr binding transcription factor TFII-I is required for efficient expression of the V␤ promoter in vivo. Mutations in the Inr element that reduced binding of TFII-I also abolished the V␤ promoter activity by ectopic TFII-I. We further biochemically identified a protease-resistant N-terminal DNA binding fragment of TFII-I, p70. When ectopically expressed, recombinant p70 bound to the V␤ Inr element with a specificity similar to that of wild-type TFII-I. More importantly, p70, which lacks independent activation functions, behaved as a dominant negative mutant that inhibited Inr-specific function of wild-type TFII-I. However, the activation functions of p70 were restored when fused to the heterologous activation domain of the yeast activator protein GAL4. Taken together, these data suggest that TFII-I functions in vivo require an intact Inr element and that the Inr-specific transcriptional functions of TFII-I are solely dictated by its N-terminal DNA binding domain and do not require its own C-terminal activation domain.

Journal of Biological Chemistry, Jun 12, 1996

The Journal of Immunology, 2007

Gene Expression Patterns, Oct 31, 2003

BEN is a member of the TFII-I family of transcription factors, characterized by the presence of m... more BEN is a member of the TFII-I family of transcription factors, characterized by the presence of multiple helix-loop-helix repeat domains. Our immunohistochemical analysis demonstrated broad and extensive expression of BEN during mouse pre- and postimplantation development, with highest levels occurring during early to midgestation. Maternally expressed BEN is present in both the cytoplasm and nuclei of the zygote; however, it retains a predominantly nuclear localization between the two-cell stage of development and early blastocyst stages. This nuclear expression is observed in most tissues throughout development. Although, it is interesting to note that at E4.5-6.5, during early gastrulation stage, BEN is localized in the cytoplasm. At later stages, BEN retains an extensive expression pattern in a variety of developing systems implicating its involvement in tissue development and organogenesis.

Molecular and Cellular Biology, 1998

In our effort to understand the transcriptional regulation of naturally occurring TATA-less but i... more In our effort to understand the transcriptional regulation of naturally occurring TATA-less but initiator (Inr)-containing genes, we have employed the murine T-cell receptor Vβ 5.2 promoter as a model. Here we show by transient-transfection assays that the Inr binding transcription factor TFII-I is required for efficient expression of the Vβ promoter in vivo. Mutations in the Inr element that reduced binding of TFII-I also abolished the Vβ promoter activity by ectopic TFII-I. We further biochemically identified a protease-resistant N-terminal DNA binding fragment of TFII-I, p70. When ectopically expressed, recombinant p70 bound to the Vβ Inr element with a specificity similar to that of wild-type TFII-I. More importantly, p70, which lacks independent activation functions, behaved as a dominant negative mutant that inhibited Inr-specific function of wild-type TFII-I. However, the activation functions of p70 were restored when fused to the heterologous activation domain of the yeast a...

Journal of Biological Chemistry

Frontiers in Human Neuroscience, 2009

Epigenetics & chromatin, 2015

Signaling via B cell receptor (BCR) and Toll-like receptors (TLRs) results in activation of B cel... more Signaling via B cell receptor (BCR) and Toll-like receptors (TLRs) results in activation of B cells with distinct physiological outcomes, but transcriptional regulatory mechanisms that drive activation and distinguish these pathways remain unknown. Two hours after ligand exposure RNA-seq, ChIP-seq and computational methods reveal that BCR- or TLR-mediated activation of primary resting B cells proceeds via a large set of shared and a smaller subset of distinct signal-selective transcriptional responses. BCR stimulation resulted in increased global recruitment of RNA Pol II to promoters that appear to transit slowly to downstream regions. Conversely, lipopolysaccharide (LPS) stimulation involved an enhanced RNA Pol II transition from initiating to elongating mode accompanied by greater H3K4me3 activation markings compared to BCR stimulation. These rapidly diverging transcriptomic landscapes also show distinct repressing (H3K27me3) histone signatures, mutually exclusive transcription f...

Trends in biochemical sciences, 2015

Early studies established that transcription initiates within an approximately 50bp DNA segment c... more Early studies established that transcription initiates within an approximately 50bp DNA segment capable of nucleating the assembly of RNA polymerase II (Pol II) and associated general transcription factors (GTFs) necessary for transcriptional initiation; this region is called a core promoter. Subsequent analyses identified a series of conserved DNA sequence elements, present in various combinations or not at all, in core promoters. Recent genome-wide analyses have provided further insights into the complexity of core promoter architecture and function. Here we review recent studies that delineate the active role of core promoters in the transcriptional regulation of diverse physiological systems.

Molecular and cellular biology, 1999

Bruton's tyrosine kinase (Btk) is required for normal B-cell development, as defects in Btk l... more Bruton's tyrosine kinase (Btk) is required for normal B-cell development, as defects in Btk lead to X-linked immunodeficiency (xid) in mice and X-linked agammaglobulinemia (XLA) in humans. Here we demonstrate a functional interaction between the multifunctional transcription factor TFII-I and Btk. Ectopic expression of wild-type Btk enhances TFII-I-mediated transcriptional activation and its tyrosine phosphorylation in transient-transfection assays. Mutation of Btk in either the PH domain (R28C, as in the murine xid mutation) or the kinase domain (K430E) compromises its ability to enhance both the tyrosine phosphorylation and the transcriptional activity of TFII-I. TFII-I associates constitutively in vivo with wild-type Btk and kinase-inactive Btk but not xid Btk. However, membrane immunoglobulin M cross-linking in B cells leads to dissociation of TFII-I from Btk. We further show that while TFII-I is found in both the nucleus and cytoplasm of wild-type and xid primary resting B ...

Molecular and cellular biology, 1998

The transcription factor TFII-I was initially isolated as a factor that can bind to initiator ele... more The transcription factor TFII-I was initially isolated as a factor that can bind to initiator elements in core promoters. Recent evidence suggests that TFII-I may also have a role in signal transduction. We have found that overexpression of TFII-I can enhance the response of the wild-type c-fos promoter to a variety of stimuli. This effect depends on the c-fos c-sis-platelet-derived growth factor-inducible factor binding element (SIE) and serum response element (SRE). There is no effect of cotransfected TFII-I on the TATA box containing the c-fos basal promoter. Three TFII-I binding sites can be found in c-fos promoter. Two of these overlap the c-fos SIE and SRE, and another is located just upstream of the TATA box. Mutations that distinguish between serum response factor (SRF), STAT, and TFII-I binding to the c-fos SIE and SRE suggest that the binding of TFII-I to these elements is important for c-fos induction in conjunction with the SRF and STAT transcription factors. Moreover, T...

Molecular Cell, 2006

Multifunctional transcription factor TFII-I has two spliced isoforms (D and b) in murine fibrobla... more Multifunctional transcription factor TFII-I has two spliced isoforms (D and b) in murine fibroblasts. Here we show that these isoforms have distinct subcellular localization and mutually exclusive transcription functions in the context of growth factor signaling. In the absence of signaling, TFII-Ib is nuclear and recruited to the c-fos promoter in vivo. But upon growth factor stimulation, the promoter recruitment is abolished and it is exported out of the nucleus. Moreover, isoform-specific silencing of TFII-Ib results in transcriptional activation of the c-fos gene. In contrast, TFII-ID is largely cytoplasmic in the resting state but translocates to the nucleus upon growth factor signaling, undergoes signal-induced recruitment to the same site on the c-fos promoter, and activates the gene. Importantly, activated TFII-ID interacts with Erk1/2 (MAPK) kinase in the cell cytoplasm and imports the Erk1/2 to the nucleus, thereby transducing growth factor signaling. Our results identify a unique growth factor signaling pathway controlled by opposing activities of two TFII-I spliced isoforms.

Frontiers in Immunology

Transcriptional regulation of cells in the immune system must be strictly controlled at multiple ... more Transcriptional regulation of cells in the immune system must be strictly controlled at multiple levels to ensure that a proper immune response is elicited only when required. Analysis in bulk, or ensemble of cells, provides a wealth of important information leading to a better understanding of the various molecular steps and mechanisms involved in regulating gene expression in immune cells. However, given the substantial heterogeneity of these cells, it is imperative now to decipher these mechanisms at a single cell level. Here I bring together several recent examples to review our understanding of transcriptional regulation of the immune system via single cell analysis and to further illustrate the immense power of such analyses to interrogate immune cell heterogeneity.

Frontiers in Immunology, Jun 14, 2019

Transcriptional regulation of cells in the immune system must be strictly controlled at multiple ... more Transcriptional regulation of cells in the immune system must be strictly controlled at multiple levels to ensure that a proper immune response is elicited only when required. Analysis in bulk, or ensemble of cells, provides a wealth of important information leading to a better understanding of the various molecular steps and mechanisms involved in regulating gene expression in immune cells. However, given the substantial heterogeneity of these cells, it is imperative now to decipher these mechanisms at a single cell level. Here I bring together several recent examples to review our understanding of transcriptional regulation of the immune system via single cell analysis and to further illustrate the immense power of such analyses to interrogate immune cell heterogeneity.

Molecular Biology of the Cell

The adult human body is composed of nearly 37 trillion cells, each with potentially unique molecu... more The adult human body is composed of nearly 37 trillion cells, each with potentially unique molecular characteristics. This Perspective describes some of the challenges and opportunities faced in mapping the molecular characteristics of these cells in specific regions of the body and highlights areas for international collaboration toward the broader goal of comprehensively mapping the human body with cellular resolution.

Science Advances

The NIH Roadmap Epigenomics Program was launched to deliver reference epigenomic data from human ... more The NIH Roadmap Epigenomics Program was launched to deliver reference epigenomic data from human tissues and cells, develop tools and methods for analyzing the epigenome, discover novel epigenetic marks, develop methods to manipulate the epigenome, and determine epigenetic contributions to diverse human diseases. Here, we comment on the outcomes from this program: the scientific contributions made possible by a consortium approach and the challenges, benefits, and lessons learned from this group science effort.

Science advances, 2018

It has become exceedingly important to understand the precise molecular profiles of the nearly 40... more It has become exceedingly important to understand the precise molecular profiles of the nearly 40 trillion cells in an adult human because of their role in determining health, disease, and therapeutic outcome. The National Institutes of Health (NIH) Common Fund-supported Single Cell Analysis Program (SCAP) was designed to address this challenge. In this review, we outline the original program goals and provide a perspective on the impact of the program as a catalyst for exploration of heterogeneity of human tissues at the cellular level. We believe that the technological advances in single-cell RNA sequencing and multiplexed imaging combined with computational methods made by this program will undoubtedly have an impact on broad and robust applications of single-cell analyses in both health and disease research.

Trends in molecular medicine, 2017

The biochemical properties of the signal-induced multifunctional transcription factor II-I (TFII-... more The biochemical properties of the signal-induced multifunctional transcription factor II-I (TFII-I) indicate that it is involved in a variety of gene regulatory processes. Although gene ablation in murine models and cell-based assays show that it is encoded by an essential gene, GTF2I/Gtf2i, its physiologic role in human disorders was relatively unknown until recently. Novel studies show that it is involved in an array of human diseases including neurocognitive disorders, systemic lupus erythematosus (SLE), and cancer. Here I bring together these diverse observations to illustrate its multiple pathophysiologic functions and further conjecture on how these could be related to its known biochemical properties. I expect that a better understanding of these 'structure-function' relationships would lead to future diagnostic and/or therapeutic potential.

Molecular and Cellular Biology, Sep 1, 1998

In our effort to understand the transcriptional regulation of naturally occurring TATA-less but i... more In our effort to understand the transcriptional regulation of naturally occurring TATA-less but initiator (Inr)-containing genes, we have employed the murine T-cell receptor V␤ 5.2 promoter as a model. Here we show by transient-transfection assays that the Inr binding transcription factor TFII-I is required for efficient expression of the V␤ promoter in vivo. Mutations in the Inr element that reduced binding of TFII-I also abolished the V␤ promoter activity by ectopic TFII-I. We further biochemically identified a protease-resistant N-terminal DNA binding fragment of TFII-I, p70. When ectopically expressed, recombinant p70 bound to the V␤ Inr element with a specificity similar to that of wild-type TFII-I. More importantly, p70, which lacks independent activation functions, behaved as a dominant negative mutant that inhibited Inr-specific function of wild-type TFII-I. However, the activation functions of p70 were restored when fused to the heterologous activation domain of the yeast activator protein GAL4. Taken together, these data suggest that TFII-I functions in vivo require an intact Inr element and that the Inr-specific transcriptional functions of TFII-I are solely dictated by its N-terminal DNA binding domain and do not require its own C-terminal activation domain.

Journal of Biological Chemistry, Jun 12, 1996

The Journal of Immunology, 2007

Gene Expression Patterns, Oct 31, 2003

BEN is a member of the TFII-I family of transcription factors, characterized by the presence of m... more BEN is a member of the TFII-I family of transcription factors, characterized by the presence of multiple helix-loop-helix repeat domains. Our immunohistochemical analysis demonstrated broad and extensive expression of BEN during mouse pre- and postimplantation development, with highest levels occurring during early to midgestation. Maternally expressed BEN is present in both the cytoplasm and nuclei of the zygote; however, it retains a predominantly nuclear localization between the two-cell stage of development and early blastocyst stages. This nuclear expression is observed in most tissues throughout development. Although, it is interesting to note that at E4.5-6.5, during early gastrulation stage, BEN is localized in the cytoplasm. At later stages, BEN retains an extensive expression pattern in a variety of developing systems implicating its involvement in tissue development and organogenesis.

Molecular and Cellular Biology, 1998

In our effort to understand the transcriptional regulation of naturally occurring TATA-less but i... more In our effort to understand the transcriptional regulation of naturally occurring TATA-less but initiator (Inr)-containing genes, we have employed the murine T-cell receptor Vβ 5.2 promoter as a model. Here we show by transient-transfection assays that the Inr binding transcription factor TFII-I is required for efficient expression of the Vβ promoter in vivo. Mutations in the Inr element that reduced binding of TFII-I also abolished the Vβ promoter activity by ectopic TFII-I. We further biochemically identified a protease-resistant N-terminal DNA binding fragment of TFII-I, p70. When ectopically expressed, recombinant p70 bound to the Vβ Inr element with a specificity similar to that of wild-type TFII-I. More importantly, p70, which lacks independent activation functions, behaved as a dominant negative mutant that inhibited Inr-specific function of wild-type TFII-I. However, the activation functions of p70 were restored when fused to the heterologous activation domain of the yeast a...

Journal of Biological Chemistry

Frontiers in Human Neuroscience, 2009

Epigenetics & chromatin, 2015

Signaling via B cell receptor (BCR) and Toll-like receptors (TLRs) results in activation of B cel... more Signaling via B cell receptor (BCR) and Toll-like receptors (TLRs) results in activation of B cells with distinct physiological outcomes, but transcriptional regulatory mechanisms that drive activation and distinguish these pathways remain unknown. Two hours after ligand exposure RNA-seq, ChIP-seq and computational methods reveal that BCR- or TLR-mediated activation of primary resting B cells proceeds via a large set of shared and a smaller subset of distinct signal-selective transcriptional responses. BCR stimulation resulted in increased global recruitment of RNA Pol II to promoters that appear to transit slowly to downstream regions. Conversely, lipopolysaccharide (LPS) stimulation involved an enhanced RNA Pol II transition from initiating to elongating mode accompanied by greater H3K4me3 activation markings compared to BCR stimulation. These rapidly diverging transcriptomic landscapes also show distinct repressing (H3K27me3) histone signatures, mutually exclusive transcription f...

Trends in biochemical sciences, 2015

Early studies established that transcription initiates within an approximately 50bp DNA segment c... more Early studies established that transcription initiates within an approximately 50bp DNA segment capable of nucleating the assembly of RNA polymerase II (Pol II) and associated general transcription factors (GTFs) necessary for transcriptional initiation; this region is called a core promoter. Subsequent analyses identified a series of conserved DNA sequence elements, present in various combinations or not at all, in core promoters. Recent genome-wide analyses have provided further insights into the complexity of core promoter architecture and function. Here we review recent studies that delineate the active role of core promoters in the transcriptional regulation of diverse physiological systems.

Molecular and cellular biology, 1999

Bruton's tyrosine kinase (Btk) is required for normal B-cell development, as defects in Btk l... more Bruton's tyrosine kinase (Btk) is required for normal B-cell development, as defects in Btk lead to X-linked immunodeficiency (xid) in mice and X-linked agammaglobulinemia (XLA) in humans. Here we demonstrate a functional interaction between the multifunctional transcription factor TFII-I and Btk. Ectopic expression of wild-type Btk enhances TFII-I-mediated transcriptional activation and its tyrosine phosphorylation in transient-transfection assays. Mutation of Btk in either the PH domain (R28C, as in the murine xid mutation) or the kinase domain (K430E) compromises its ability to enhance both the tyrosine phosphorylation and the transcriptional activity of TFII-I. TFII-I associates constitutively in vivo with wild-type Btk and kinase-inactive Btk but not xid Btk. However, membrane immunoglobulin M cross-linking in B cells leads to dissociation of TFII-I from Btk. We further show that while TFII-I is found in both the nucleus and cytoplasm of wild-type and xid primary resting B ...

Molecular and cellular biology, 1998

The transcription factor TFII-I was initially isolated as a factor that can bind to initiator ele... more The transcription factor TFII-I was initially isolated as a factor that can bind to initiator elements in core promoters. Recent evidence suggests that TFII-I may also have a role in signal transduction. We have found that overexpression of TFII-I can enhance the response of the wild-type c-fos promoter to a variety of stimuli. This effect depends on the c-fos c-sis-platelet-derived growth factor-inducible factor binding element (SIE) and serum response element (SRE). There is no effect of cotransfected TFII-I on the TATA box containing the c-fos basal promoter. Three TFII-I binding sites can be found in c-fos promoter. Two of these overlap the c-fos SIE and SRE, and another is located just upstream of the TATA box. Mutations that distinguish between serum response factor (SRF), STAT, and TFII-I binding to the c-fos SIE and SRE suggest that the binding of TFII-I to these elements is important for c-fos induction in conjunction with the SRF and STAT transcription factors. Moreover, T...

Molecular Cell, 2006

Multifunctional transcription factor TFII-I has two spliced isoforms (D and b) in murine fibrobla... more Multifunctional transcription factor TFII-I has two spliced isoforms (D and b) in murine fibroblasts. Here we show that these isoforms have distinct subcellular localization and mutually exclusive transcription functions in the context of growth factor signaling. In the absence of signaling, TFII-Ib is nuclear and recruited to the c-fos promoter in vivo. But upon growth factor stimulation, the promoter recruitment is abolished and it is exported out of the nucleus. Moreover, isoform-specific silencing of TFII-Ib results in transcriptional activation of the c-fos gene. In contrast, TFII-ID is largely cytoplasmic in the resting state but translocates to the nucleus upon growth factor signaling, undergoes signal-induced recruitment to the same site on the c-fos promoter, and activates the gene. Importantly, activated TFII-ID interacts with Erk1/2 (MAPK) kinase in the cell cytoplasm and imports the Erk1/2 to the nucleus, thereby transducing growth factor signaling. Our results identify a unique growth factor signaling pathway controlled by opposing activities of two TFII-I spliced isoforms.