In vivo complex haploinsufficiency-based genetic analysis identifies a transcription factor circuit regulating Candida albicans oropharyngeal infection and epithelial cell endocytosis

Oropharyngeal candidiasis (OPC) is a common infection that complicates a wide range of medical conditions which can cause either mild or severe disease depending on the patient. The pathobiology of OPC shares many features with candidal biofilms of abiotic surfaces. The transcriptional regulation of C. albicans formation of biofilms on abiotic surfaces has been extensively characterized and involves six key transcription factors (Efg1, Ndt80, Rob1, Bcr1, Brg1, and Tec1). To determine whether this same in vitro biofilm transcriptional regulatory network played a role in OPC, we have carried out the first systematic genetic interaction analysis in a mouse model of C. albicans infection. Whereas all six transcription factors are required for in vitro biofilm formation, only three homozygous deletion mutants ( tec1 ??, bcr1 ??, and rob1 ??) and one heterozygous mutant ( tec1 ?/ TEC1 ) have reduced infectivity in a mouse model of OPC, indicating the network is more robust in vivo than in vitro. Although single mutants (heterozygous or homozygous) of BRG1 and EFG1 have no effect on fungal burden, the double heterozygous and homozygous mutants have dramatically reduced infectivity, indicating a critical genetic interaction between these two transcription factors. Using epistasis analysis, we have formulated a genetic circuit [ EFG1 + BRG1 ]→ TEC1 → BCR1 that is required for OPC infectivity and oral epithelial cell endocytosis. Surprisingly, we also found transcription factor mutants with in vitro defects in filamentation such as efg1 ?? and brg1 ?? filament during oral infection and that decreased filamentation did not correlate with decreased infectivity. Taken together, these data indicate that key in vitro biofilm transcription factors are involved in OPC but that the network characteristics and functional connections are remodeled significantly during interactions with tissues.

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Subnuclear compartmentalization of sequence-specific transcription factors and regulation of eukaryotic gene expression

Biochemistry and Cell Biology ◽

10.1139/o05-062 ◽

2005 ◽

Vol 83 (4) ◽

pp. 535-547 ◽

Cited By ~ 7

Author(s):

Gareth N Corry ◽

D Alan Underhill

Keyword(s):

Gene Expression ◽

Transcription Factor ◽

Transcription Factors ◽

Transcriptional Activation ◽

Current Knowledge ◽

Nuclear Architecture ◽

Subnuclear Localization ◽

Modular Structures

To date, the majority of the research regarding eukaryotic transcription factors has focused on characterizing their function primarily through in vitro methods. These studies have revealed that transcription factors are essentially modular structures, containing separate regions that participate in such activities as DNA binding, protein–protein interaction, and transcriptional activation or repression. To fully comprehend the behavior of a given transcription factor, however, these domains must be analyzed in the context of the entire protein, and in certain cases the context of a multiprotein complex. Furthermore, it must be appreciated that transcription factors function in the nucleus, where they must contend with a variety of factors, including the nuclear architecture, chromatin domains, chromosome territories, and cell-cycle-associated processes. Recent examinations of transcription factors in the nucleus have clarified the behavior of these proteins in vivo and have increased our understanding of how gene expression is regulated in eukaryotes. Here, we review the current knowledge regarding sequence-specific transcription factor compartmentalization within the nucleus and discuss its impact on the regulation of such processes as activation or repression of gene expression and interaction with coregulatory factors.Key words: transcription, subnuclear localization, chromatin, gene expression, nuclear architecture.

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The SCL Transcription Factor Supports Erythroid Cell Survival and Differentiation Downstream of the Erythropoietin Receptor.

Blood ◽

10.1182/blood.v104.11.818.818 ◽

2004 ◽

Vol 104 (11) ◽

pp. 818-818

Author(s):

Rachid Lahlil ◽

Richard Martin ◽

Peter D. Aplan ◽

C. Glenn Begley ◽

Jacqueline E. Damen ◽

...

Keyword(s):

Transcription Factor ◽

Cell Survival ◽

Cell Fate ◽

Genetic Interaction ◽

Fetal Liver ◽

Erythropoietin Receptor ◽

Erythroid Cell ◽

Loss Of Function

Abstract Erythroid cell development critically depends on the SCL/Tal1 transcription factor and on erythropoietin signalling. In the present study, we have taken several approaches to show that the two genes operate within the same pathway to consolidate the erythroid lineage. Signaling through the erythropoietin receptor (EpoR) upregulates SCL protein levels in a clonal cell line (TF-1) in vitro, and in murine fetal liver cells in vivo, when Epor−/− cells were compared to those of wild type littermates at E12.5. In addition, we provide functional evidence for a linear pathway from EpoR to SCL that regulates erythropoiesis. Interfering with SCL induction or SCL function prevents the anti-apoptotic effect of Epo in TF-1 cells and conversely, ectopic SCL expression is sufficient to substitute for Epo to transiently maintain cell survival. In vivo, SCL gain of function complements the cellular defects in Epor−/− embryos to support cell survival and maturation during primitive and definitive erythropoiesis, as assessed by cellular and histological analyses of Epor−/− SCLtg embryos. Moreover, several erythroid specific genes that are decreased in Epor−/− embryos are rescued by the SCL transgene including glycophorinA, bH1 and bmaj globin, providing molecular confirmation of the functional and genetic interaction between Epor and SCL. Conversely, erythropoiesis becomes deficient in compound Epor+/−SCL+/− heterozygote mice, indicating that the genetic interaction between EpoR and SCL is synthetic. Finally, using EpoR mutants that harbour well defined signalling deficiencies, combined with gain and loss of function approaches for specific kinases, we identify MAPK as the major signal transduction pathway downstream of EpoR that upregulates SCL function, necessary for erythroid cell survival and differentiation. Taken together, our observations are consistent with the view that cytokines can influence cell fate by altering the dosage of lineage transcriptional regulators.

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Overlapping and Unique Roles for C-Terminal Binding Protein 1 (CtBP1) and CtBP2 during Mouse Development

Molecular and Cellular Biology ◽

10.1128/mcb.22.15.5296-5307.2002 ◽

2002 ◽

Vol 22 (15) ◽

pp. 5296-5307 ◽

Cited By ~ 199

Author(s):

Jeffrey D. Hildebrand ◽

Philippe Soriano

Keyword(s):

Gene Expression ◽

Transcription Factors ◽

Genetic Interaction ◽

Binding Protein ◽

Biological Processes ◽

Vertebrate Development ◽

Mouse Development ◽

Axial Patterning ◽

Wide Range

ABSTRACT The C-terminal binding protein (CtBP) family of proteins has been linked to multiple biological processes through their association with numerous transcription factors. We generated mice harboring mutations in both Ctbp1 and Ctbp2 to address the in vivo function of CtBPs during vertebrate development. Ctbp1 mutant mice are small but viable and fertile, whereas Ctbp2-null mice show defects in axial patterning and die by E10.5 due to aberrant extraembryonic development. Mice harboring various combinations of Ctbp1 and Ctbp2 mutant alleles exhibit dosage-sensitive defects in a wide range of developmental processes. The strong genetic interaction, as well as transcription assays with CtBP-deficient cells, indicates that CtBPs have overlapping roles in regulating gene expression. We suggest that the observed phenotypes reflect the large number of transcription factors whose activities are compromised in the absence of CtBP.

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Regulation of E2F1-Dependent Gene Transcription and Apoptosis by the ETS-Related Transcription Factor GABPγ1

Molecular and Cellular Biology ◽

10.1128/mcb.22.7.2147-2158.2002 ◽

2002 ◽

Vol 22 (7) ◽

pp. 2147-2158 ◽

Cited By ~ 18

Author(s):

Ludger Hauck ◽

Rudolf G. Kaba ◽

Martin Lipp ◽

Rainer Dietz ◽

Rüdiger von Harsdorf

Keyword(s):

Transcription Factor ◽

Transcription Factors ◽

Cell Fate ◽

Binding Domain ◽

Pocket Proteins ◽

Related Transcription Factor ◽

Terminal Amino ◽

Fate Decision

ABSTRACT The E2F family of transcription factors comprises six related members which are involved in the control of the coordinated progression through the G1/S-phase transition of cell cycle or in cell fate decision. Their activity is regulated by pocket proteins, including pRb, p107, and p130. Here we show that E2F1 directly interacts with the ETS-related transcription factor GABPγ1 in vitro and in vivo. The binding domain interacting with GABPγ1 was mapped to the C-terminal amino acids 310 to 437 of E2F1, which include its transactivation and pRb binding domain. Among the E2F family of transcription factors, the interaction with GABPγ1 is restricted to E2F1. DNA-binding E2F1 complexes containing GABPγ1 are characterized by enhanced E2F1-dependent transcriptional activity. Moreover, GABPγ1 suppresses E2F1-dependent apoptosis by mechanisms other than the inhibition of the transactivation capacity of E2F1. In summary, our results provide evidence for a novel pRb-independent mechanism regulating E2F1-dependent transcription and apoptosis.

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Single-molecule imaging reveals the interplay between transcription factors, nucleosomes, and transcriptional bursting

10.1101/404681 ◽

2018 ◽

Cited By ~ 1

Author(s):

Benjamin T. Donovan ◽

Anh Huynh ◽

David A. Ball ◽

Michael G. Poirier ◽

Daniel R. Larson ◽

...

Keyword(s):

Transcription Factor ◽

Transcription Factors ◽

Single Molecule ◽

Target Genes ◽

Burst Size ◽

Yeast Cells ◽

Single Molecule Imaging ◽

Transcriptional Bursting

SummaryTranscription factors show rapid and reversible binding to chromatin in living cells, and transcription occurs in sporadic bursts, but how these phenomena are related is unknown. Using a combination of in vitro and in vivo single-molecule imaging approaches, we directly correlated binding of the transcription factor Gal4 with the transcriptional bursting kinetics of the Gal4 target genes GAL3 and GAL10 in living yeast cells. We find that Gal4 dwell times sets the transcriptional burst size. Gal4 dwell time depends on the affinity of the binding site and is reduced by orders of magnitude by nucleosomes. Using a novel imaging platform, we simultaneously tracked transcription factor binding and transcription at one locus, revealing the timing and correlation between Gal4 binding and transcription. Collectively, our data support a model where multiple polymerases initiate during a burst as long as the transcription factor is bound to DNA, and a burst terminates upon transcription factor dissociation.

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Determining Aspergillus fumigatus transcription factor expression and function during invasion of the mammalian lung

PLoS Pathogens ◽

10.1371/journal.ppat.1009235 ◽

2021 ◽

Vol 17 (3) ◽

pp. e1009235

Author(s):

Hong Liu ◽

Wenjie Xu ◽

Vincent M. Bruno ◽

Quynh T. Phan ◽

Norma V. Solis ◽

...

Keyword(s):

Transcription Factor ◽

Transcription Factors ◽

Aspergillus Fumigatus ◽

Transcriptional Profiling ◽

Transcriptional Response ◽

Gene Clusters ◽

Transcription Factor Genes ◽

And Function

To gain a better understanding of the transcriptional response of Aspergillus fumigatus during invasive pulmonary infection, we used a NanoString nCounter to assess the transcript levels of 467 A. fumigatus genes during growth in the lungs of immunosuppressed mice. These genes included ones known to respond to diverse environmental conditions and those encoding most transcription factors in the A. fumigatus genome. We found that invasive growth in vivo induces a unique transcriptional profile as the organism responds to nutrient limitation and attack by host phagocytes. This in vivo transcriptional response is largely mimicked by in vitro growth in Aspergillus minimal medium that is deficient in nitrogen, iron, and/or zinc. From the transcriptional profiling data, we selected 9 transcription factor genes that were either highly expressed or strongly up-regulated during in vivo growth. Deletion mutants were constructed for each of these genes and assessed for virulence in mice. Two transcription factor genes were found to be required for maximal virulence. One was rlmA, which is required for the organism to achieve maximal fungal burden in the lung. The other was sltA, which regulates of the expression of multiple secondary metabolite gene clusters and mycotoxin genes independently of laeA. Using deletion and overexpression mutants, we determined that the attenuated virulence of the ΔsltA mutant is due in part to decreased expression aspf1, which specifies a ribotoxin, but is not mediated by reduced expression of the fumigaclavine gene cluster or the fumagillin-pseruotin supercluster. Thus, in vivo transcriptional profiling focused on transcription factors genes provides a facile approach to identifying novel virulence regulators.

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CUX1—Transcriptional Master Regulator of Tumor Progression in Pancreatic Neuroendocrine Tumors

Cancers ◽

10.3390/cancers12071957 ◽

2020 ◽

Vol 12 (7) ◽

pp. 1957

Author(s):

Sebastian Krug ◽

Julia Weissbach ◽

Annika Blank ◽

Aurel Perren ◽

Johannes Haybaeck ◽

...

Keyword(s):

Transcription Factor ◽

Mouse Model ◽

Neuroendocrine Tumours ◽

Tumour Progression ◽

Free Survival ◽

Treatment Naïve ◽

Pancreatic Neuroendocrine Tumours ◽

The Impact

Recently, we identified the homeodomain transcription factor Cut homeobox 1 (CUX1) as mediator of tumour de-differentiation and metastatic behaviour in human insulinoma patients. In insulinomas, CUX1 enhanced tumour progression by stimulating proliferation and angiogenesis in vitro and in vivo. In patients with non-functional pancreatic neuroendocrine tumours (PanNET), however, the impact of CUX1 remains to be elucidated. Here, we analysed CUX1 expression in two large independent cohorts (n = 43 and n = 141 tissues) of non-functional treatment-naïve and pre-treated PanNET patients, as well as in the RIP1Tag2 mouse model of pancreatic neuroendocrine tumours. To further assess the functional role of CUX1, expression profiling of DNA damage-, proliferation- and apoptosis-associated genes was performed in CUX1-overexpressing Bon-1 cells. Validation of differentially regulated genes was performed in Bon-1 and QGP1 cells with knock-down and overexpression strategies. CUX1 expression assessed by a predefined immunoreactivity score (IRS) was significantly associated with shorter progression-free survival (PFS) of pre-treated PanNET patients (23 vs. 8 months; p = 0.005). In treatment-naïve patients, CUX1 was negatively correlated with grading and recurrence-free survival (mRFS of 39 versus 8 months; p = 0.022). In both groups, high CUX1 levels indicated a metastatic phenotype. Functionally, CUX1 upregulated expression of caspases and death associated protein kinase 1 (DAPK1), known as mediators of tumour progression and resistance to cytotoxic drugs. This was also confirmed in both cell lines and human tissues. In the RIP1Tag2 mouse model, CUX1 expression was associated with advanced tumour stage and resistance to apoptosis. In summary, we identified the transcription factor CUX1 as mediator of tumour progression in non-functional PanNET in vitro and in vivo, indicating that the CUX1-dependent signalling network is a promising target for future therapeutic intervention.

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GATA-1 Forms Distinct Activating and Repressive Complexes in Erythroid Cells.

Blood ◽

10.1182/blood.v104.11.356.356 ◽

2004 ◽

Vol 104 (11) ◽

pp. 356-356

Author(s):

John Strouboulis ◽

Patrick Rodriguez ◽

Edgar Bonte ◽

Jeroen Krijgsveld ◽

Katarzyna Kolodziej ◽

...

Keyword(s):

Transcription Factor ◽

Transcription Factors ◽

Chromatin Remodeling ◽

Gene Activation ◽

Erythroid Differentiation ◽

Specific Gene ◽

Erythroid Cells ◽

In Erythroid Cells

Abstract GATA-1 is a key transcription factor essential for the differentiation of the erythroid, megakaryocytic and eosinophilic lineages. GATA-1 functions in erythropoiesis involve lineage-specific gene activation and repression of early hematopoietic transcription programs. GATA-1 is known to interact with other transcription factors, such as FOG-1, TAL-1 and Sp1 and also with CBP/p300 and the SWI/SNF chromatin remodeling complex in vitro. Despite this information the molecular basis of its essential functions in erythropoiesis remains unclear. We show here that GATA-1 is mostly present in a high (> 670kDa) molecular weight complex that appears to be dynamic during erythroid differentiation. In order to characterize the GATA-1 complex(es) from erythroid cells, we employed an in vivo biotinylation tagging approach in mouse erythroleukemic (MEL) cells1. Briefly, this involved the fusion of a small (23aa) peptide tag to GATA-1 and its specific, efficient biotinylation by the bacterial BirA biotin ligase which is co-expressed with tagged GATA-1 in MEL cells. Nuclear extracts expressing biotinylated tagged GATA-1 were bound directly to streptavidin beads and co-purifying proteins were identified by mass spectrometry. In addition to the known GATA-1-interacting transcription factors FOG-1, TAL-1 and Ldb-1, we describe novel interactions with the essential hematopoietic transcription factor Gfi-1b and the chromatin remodeling complexes MeCP1 and ACF/WCRF. Significantly, GATA-1 interaction with the repressive MeCP1 complex requires FOG-1. We also show in erythroid cells that GATA-1, FOG-1 and MeCP1 are stably bound to repressed genes representing early hematopoietic (e.g. GATA-2) or alternative lineage-specific (e.g. eosinophilic) transcription programs, whereas the GATA-1/Gfi1b complex is bound to repressed genes involved in cell proliferation. In contrast, GATA-1 and TAL-1 are bound to the active erythroid-specific EKLF gene. Our findings on GATA-1 complexes provide novel insight as to the critical roles that GATA-1 plays in many aspects of erythropoiesis by revealing the GATA-1 partners in the execution of specific functions.

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Identification of Key Cellular Regulators That Interact with Id1 To Control Hematopoietic Stem Cell Behavior.

Blood ◽

10.1182/blood.v108.11.1319.1319 ◽

2006 ◽

Vol 108 (11) ◽

pp. 1319-1319

Author(s):

Vladimir Jankovic ◽

Alessia Ciarrocchi ◽

Tony DeBlasio ◽

Robert Benezra ◽

Stephen D. Nimer

Keyword(s):

Transcription Factors ◽

Transcriptional Repression ◽

Genetic Interaction ◽

Dominant Negative ◽

Double Knockout ◽

Self Renewal ◽

Hematopoietic Stem ◽

Helix Loop Helix

Abstract The ability of hematopoietic stem cells to tightly regulate the transition from relative quiescence and self-renewal to the transiently amplifying, differentiating progenitor fate is critical for HSC homeostasis as well as their regenerative capacity. We have recently described the diminished frequency and rapid exhaustion of HSC self-renewal capacity in the absence of the dominant negative helix-loop-helix molecule Id1. Furthermore, Id1 null HSCs have an increased rate of cycling, coupled with accelerated myeloid commitment both in vivo and in vitro. This is reflected in the elevated expression of myelo-erythroid transcription factors (c/EBPalpha and GATA1) within the Lin−c-kit+Sca-1+ population - “myeloid priming”. The major targets of Id1 mediated transcriptional repression are the ubiquitous E protein E2A as well as Ets transcription factors (Ets1 and Ets2). We hypothesized that the unrestrained activity of these and/or other targets of Id1 transcriptional repression leads to premature HSC commitment in Id1 null animals. Indeed, we show that HSC differentiation in culture can be delayed by transduction of E2A directed shRNA specifically in Id1 null, but not in wild-type Id1 expressing cells. This indicates an abnormal E2A activity in Id1 null HSCs that could be responsible for their increased differentiation status. To further define the transcriptional deregulation in Id1 null HSCs, we have used the Affymetrix microarray technology. We observed ~3 fold increased expression of the CDK inhibitor p21 in freshly isolated Id1 null HSCs and have confirmed this result by multiple independent qPCR measurements. The transcriptional induction of p21 by E2A as well as its repression by Id1 have been well established. Therefore, the observed p21 induction could be explained by the elevated level of E2A activity in HSCs in the absence of Id1 expression. To explore the functional significance of Id1 mediated p21 regulation in HSCs, we have generated p21/Id1 double knockout animals. Surprisingly, despite its reported function in restricting the cell cycle entry of normal HSCs, we show that in the context of Id1 loss, p21 expression is required for the accelerated HSC cycling, and unlike Id1 single null HSCs, p21/Id1 double knockout HSCs do not show accelerated myeloid differentiation in culture. Therefore, we propose that Id1 actively represses E2A activity in HSCs, as well as the induction of p21, which could be an important component of the HSC commitment program. Further studies will be presented defining the in vivo relevance of the Id1/p21 genetic interaction for HSC growth and differentiation.

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LEAFY is a pioneer transcription factor and licenses cell reprogramming to floral fate

Nature Communications ◽

10.1038/s41467-020-20883-w ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Run Jin ◽

Samantha Klasfeld ◽

Yang Zhu ◽

Meilin Fernandez Garcia ◽

Jun Xiao ◽

...

Keyword(s):

Transcription Factor ◽

Transcription Factors ◽

Cell Fate ◽

Chromatin Accessibility ◽

Binding Motifs ◽

Chromatin Remodelers ◽

Pioneer Transcription Factor ◽

Chromatin Reprogramming

AbstractMaster transcription factors reprogram cell fate in multicellular eukaryotes. Pioneer transcription factors have prominent roles in this process because of their ability to contact their cognate binding motifs in closed chromatin. Reprogramming is pervasive in plants, whose development is plastic and tuned by the environment, yet little is known about pioneer transcription factors in this kingdom. Here, we show that the master transcription factor LEAFY (LFY), which promotes floral fate through upregulation of the floral commitment factor APETALA1 (AP1), is a pioneer transcription factor. In vitro, LFY binds to the endogenous AP1 target locus DNA assembled into a nucleosome. In vivo, LFY associates with nucleosome occupied binding sites at the majority of its target loci, including AP1. Upon binding, LFY ‘unlocks’ chromatin locally by displacing the H1 linker histone and by recruiting SWI/SNF chromatin remodelers, but broad changes in chromatin accessibility occur later. Our study provides a mechanistic framework for patterning of inflorescence architecture and uncovers striking similarities between LFY and animal pioneer transcription factor.

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