biswanath chatterjee - Academia.edu (original) (raw)

Papers by biswanath chatterjee

A causal relationship exists among the aging process, organ decay and dis-function, and the occur... more A causal relationship exists among the aging process, organ decay and dis-function, and the occurrence of various diseases including cancer. A genetically engineered mouse model, termed EklfK74R/K74R or Eklf(K74R), carrying mutation on the well-conserved sumoylation site of the hematopoietic transcription factor KLF1/ EKLF has been generated that possesses extended lifespan and healthy characteristics including cancer resistance. We show that the healthy longevity characteristics of the Eklf(K74R) mice, as exemplified by their higher anti-cancer capability, are likely gender-, age- and genetic background-independent. Significantly, the anti-cancer capability, in particular that against melanoma as well as hepatocellular carcinoma, and lifespan-extending property of Eklf(K74R) mice could be transferred to wild-type mice via transplantation of their bone marrow mononuclear cells at young age of the latter. Furthermore, NK(K74R) cells carry higher in vitro cancer cell-killing ability t...

<p>(A) Amyloidogenesis of mPrP(23–230) (22 µM) in 1 M GdnHCl, 3 M urea in PBS, pH 6.0, incu... more <p>(A) Amyloidogenesis of mPrP(23–230) (22 µM) in 1 M GdnHCl, 3 M urea in PBS, pH 6.0, incubated at 37°C with vigorous shaking in the absence of seed (denoted by closed, half-filled and open squares for three independent measurements) or in the presence of 20 µL (denoted by closed, half-filled and open circles for three independent measurements) or 40 µL (denoted by closed, half-filled and open up triangles for three independent measurements) of sonicated mPrP(23–230) seed. In these three seeding experiments, mPrP(23–230) seed was prepared independently and three batches of seed contained 88.7, 88.3 and 96.2 pmoles of mPrP(23–230) monomer per microliter seed solution. (B) Amyloidogenesis of mPrP(23–230) under the same conditions as in (A) cross-seeded with 20 µL of sonicated mPrP(107–143) seed (denoted by closed, half-filled and open circles for three independent measurements). The seed contained 131.9, 129.1, and 119.7 pmoles of mPrP(107–143) per microliter solution in these three independent measurements. The data of spontaneous amyloidogenesis without seeding are also shown in the same plot for comparison.</p

<p>mPrP(107–143) (50 µM) in 140 mM NaCl and 20 mM NaOAc, pH 3.7, was incubated at 25°C with... more <p>mPrP(107–143) (50 µM) in 140 mM NaCl and 20 mM NaOAc, pH 3.7, was incubated at 25°C without shaking alone (denoted by closed, half-filled and open squares for three independent measurements), in the presence of 30 µL of mPrP(23–230) seed (denoted by closed, half-filled and open circles for three independent measurements), or in the presence of the same amount of mPrP(23–230) seed digested by proteinase K (denoted by closed, half-filled and open up triangles for three independent measurements).</p

Skeletal muscle, 2016

Master transcription factor MyoD can initiate the entire myogenic gene expression program which d... more Master transcription factor MyoD can initiate the entire myogenic gene expression program which differentiates proliferating myoblasts into multinucleated myotubes. We previously demonstrated that histone methyltransferase KMT1A associates with and inhibits MyoD in proliferating myoblasts, and must be removed to allow differentiation to proceed. It is known that pro-myogenic signaling pathways such as PI3K/AKT and p38α MAPK play critical roles in enforcing associations between MyoD and transcriptional activators, while removing repressors. However, the mechanism which displaces KMT1A from MyoD, and the signals responsible, remain unknown. To investigate the role of p38α on MyoD-mediated differentiation, we utilized C2C12 myoblast cells as an in vitro model. p38α activity was either augmented via overexpression of a constitutively active upstream kinase or blocked via lentiviral delivery of a specific p38α shRNA or treatment with p38α/β inhibitor SB203580. Overexpression of KMT1A in ...

Acta neuropathologica, Jan 12, 2016

For proper mammalian brain development and functioning, the translation of many neuronal mRNAs ne... more For proper mammalian brain development and functioning, the translation of many neuronal mRNAs needs to be repressed without neuronal activity stimulations. We have discovered that the expression of a subclass of neuronal proteins essential for neurodevelopment and neuron plasticity is co-regulated at the translational level by TDP-43 and the Fragile X Syndrome protein FMRP. Using molecular, cellular and imaging approaches, we show that these two RNA-binding proteins (RBP) co-repress the translation initiation of Rac1, Map1b and GluR1 mRNAs, and consequently the hippocampal spinogenesis. The co-repression occurs through binding of TDP-43 to mRNA(s) at specific UG/GU sequences and recruitment of the inhibitory CYFIP1-FMRP complex by its glycine-rich domain. This novel regulatory scenario could be utilized to silence a significant portion of around 160 common target mRNAs of the two RBPs. The study establishes a functional/physical partnership between FMRP and TDP-43 that mechanistica...

Knockdown of p38α was monitored by western blot analysis of cell extracts from primary HsMB cells... more Knockdown of p38α was monitored by western blot analysis of cell extracts from primary HsMB cells expressing control scramble shRNA (Ctrl) or p38α shRNA, probed with antibodies for p38α, total p38, β-actin as loading control. Decreased levels of both p38α and total p38 were observed by p38α shRNA relative to Ctrl shRNA. (PPTX 47 kb)

Western blot analysis verified phosphorylated KMT1A by re-probing the membrane of the western blo... more Western blot analysis verified phosphorylated KMT1A by re-probing the membrane of the western blot results presented in Fig. 4a with a separate KMT1A antibody. (B) Western blot analysis of C2C12 cells expressing control scramble shRNA (Ctrl) or KMT1A shRNA via lentiviral delivery grown in GM or DM, probed with antibodies against KMT1A, and β-actin as loading control. Decreased levels of both under- and phosphorylated KMT1A were observed by KMT1A shRNA relative to Ctrl. (C) Flag-KMT1A overexpression was monitored by western blot analysis of cell extracts following it expression in 293A cells (293A-KMT1A-F) via lentiviral delivery. (D) In vitro kinase assays was performed for p38α activity using GST or GST-ATF2 as substrate by in vitro kinase assays in the presence and absence of SB. Commassie and autoradiography detected inputs GST/GST-ATF2 proteins and phosphorylated ATF2, respectively. (PPTX 99 kb)

(A) Ectopic Flag-KMT1A expression was determined by western blot analysis of C2-4RE-luc and C2-4R... more (A) Ectopic Flag-KMT1A expression was determined by western blot analysis of C2-4RE-luc and C2-4RE-Luc/KMT1A-F cells grown in GM, probed with antibodies to Flag-KMT1A, total KMT1A, MyoD, and β-actin as loading control. Flag-KMT1A expression was observed only in C2-4RE-luc/KMT1A-F cells. (B) Luciferase activity was monitored in C2-4RE-luc cells expressing vector control (Em), MKK6EE or MMK6DN grown in GM and values expressed after protein normalization as fold activation. Error bar, ±SEM (n = 3). MyoD-responsive reporter luciferase gene activation was observed by MKK6EE but not MKKDN in these cells. (PPTX 59 kb)

The principal event underlying the development of prion disease is the conversion of soluble cell... more The principal event underlying the development of prion disease is the conversion of soluble cellular prion protein (PrPC) into its disease-causing isoform, PrPSc. This conversion is associated with a marked change in secondary structure from predominantly a-helical to a high b-sheet content, ultimately leading to the formation of aggregates consisting of ordered fibrillar assemblies referred to as amyloid. In vitro, recombinant prion proteins and short prion peptides from various species have been shown to form amyloid under various conditions and it has been proposed that, theoretically, any protein and peptide could form amyloid under appropriate conditions. To identify the peptide segment involved in the amyloid core formed from recombinant full-length mouse prion protein mPrP(23–230), we carried out seed-induced amyloid formation from recombinant prion protein in the presence of seeds generated from the short prion peptides mPrP(107–143), mPrP(107–126), and mPrP(127–143). Our r...

Expression of HA-tagged MKK6EE and MKK6DN in C2C12 cells was verified by western blot analysis, p... more Expression of HA-tagged MKK6EE and MKK6DN in C2C12 cells was verified by western blot analysis, probed with antibodies against HA, and β-actin as loading control. (B) Control IgG or anti-phospho-p38 immunoprecipitates retrieved from extracts of indicated cells grown in GM or DM were subjected to in vitro kinase assays to monitor p38 activation using GST-ATF2 as substrate. Autoradiography and Commassie detected phosphorylated ATF2 and GST-ATF2 protein, respectively. (PPTX 75 kb)

Cell Reports

Highlights d TDP-43 cooperates with FMRP to modulate anterograde transport of dendritic mRNP d TD... more Highlights d TDP-43 cooperates with FMRP to modulate anterograde transport of dendritic mRNP d TDP-43 and Staufen1 co-regulate retrograde transport of mRNP in neuronal dendrites d TDP-43 regulates kinetics and dynamics of coupled dendritic transport-translation d TDP-43 regulates spine base-to-spine transport of Rac1 mRNA for local translation Authors

Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanisms

The DNA methylation program in vertebrates is an essential part of the epigenetic regulatory casc... more The DNA methylation program in vertebrates is an essential part of the epigenetic regulatory cascade of development, cell differentiation, and progression of diseases including cancer. While the DNA methyltransferases (DNMTs) are responsible for the in vivo conversion of cytosine (C) to methylated cytosine (5mC), demethylation of 5mC on cellular DNA could be accomplished by the combined action of the ten-eleven translocation (TET) enzymes and DNA repair. Surprisingly, the mammalian DNMTs also possess active DNA demethylation activity in vitro in a Ca2+- and redox conditions-dependent manner, although little is known about its molecular mechanisms and occurrence in a cellular context. In this study, we have used LC-MS/MS to track down the fate of the methyl group removed from 5mC on DNA by mouse DNMT3B in vitro and found that it becomes covalently linked to the DNA methylation catalytic cysteine of the enzyme. We also show that Ca2+ homeostasis-dependent but TET1/TET2/TET3/TDG-independent demethylation of methylated episomal DNA by mouse DNMT3A or DNMT3B can occur in transfected human HEK 293 and mouse embryonic stem (ES) cells. Based on these results, we present a tentative working model of Ca2+ and redox conditions-dependent active DNA demethylation by DNMTs. Our study substantiates the potential roles of the vertebrate DNMTs as double-edged swords in DNA methylation-demethylation during Ca2+-dependent physiological processes.

International Journal of Molecular Sciences

The intrinsic cellular heterogeneity and molecular complexity of the mammalian nervous system rel... more The intrinsic cellular heterogeneity and molecular complexity of the mammalian nervous system relies substantially on the dynamic nature and spatiotemporal patterning of gene expression. These features of gene expression are achieved in part through mechanisms involving various epigenetic processes such as DNA methylation, post-translational histone modifications, and non-coding RNA activity, amongst others. In concert, another regulatory layer by which RNA bases and sugar residues are chemically modified enhances neuronal transcriptome complexity. Similar RNA modifications in other systems collectively constitute the cellular epitranscriptome that integrates and impacts various physiological processes. The epitranscriptome is dynamic and is reshaped constantly to regulate vital processes such as development, differentiation and stress responses. Perturbations of the epitranscriptome can lead to various pathogenic conditions, including cancer, cardiovascular abnormalities and neurol...

A causal relationship exists among the aging process, organ decay and dis-function, and the occur... more A causal relationship exists among the aging process, organ decay and dis-function, and the occurrence of various diseases including cancer. A genetically engineered mouse model, termed EklfK74R/K74R or Eklf(K74R), carrying mutation on the well-conserved sumoylation site of the hematopoietic transcription factor KLF1/ EKLF has been generated that possesses extended lifespan and healthy characteristics including cancer resistance. We show that the healthy longevity characteristics of the Eklf(K74R) mice, as exemplified by their higher anti-cancer capability, are likely gender-, age- and genetic background-independent. Significantly, the anti-cancer capability, in particular that against melanoma as well as hepatocellular carcinoma, and lifespan-extending property of Eklf(K74R) mice could be transferred to wild-type mice via transplantation of their bone marrow mononuclear cells at young age of the latter. Furthermore, NK(K74R) cells carry higher in vitro cancer cell-killing ability t...

<p>(A) Amyloidogenesis of mPrP(23–230) (22 µM) in 1 M GdnHCl, 3 M urea in PBS, pH 6.0, incu... more <p>(A) Amyloidogenesis of mPrP(23–230) (22 µM) in 1 M GdnHCl, 3 M urea in PBS, pH 6.0, incubated at 37°C with vigorous shaking in the absence of seed (denoted by closed, half-filled and open squares for three independent measurements) or in the presence of 20 µL (denoted by closed, half-filled and open circles for three independent measurements) or 40 µL (denoted by closed, half-filled and open up triangles for three independent measurements) of sonicated mPrP(23–230) seed. In these three seeding experiments, mPrP(23–230) seed was prepared independently and three batches of seed contained 88.7, 88.3 and 96.2 pmoles of mPrP(23–230) monomer per microliter seed solution. (B) Amyloidogenesis of mPrP(23–230) under the same conditions as in (A) cross-seeded with 20 µL of sonicated mPrP(107–143) seed (denoted by closed, half-filled and open circles for three independent measurements). The seed contained 131.9, 129.1, and 119.7 pmoles of mPrP(107–143) per microliter solution in these three independent measurements. The data of spontaneous amyloidogenesis without seeding are also shown in the same plot for comparison.</p

<p>mPrP(107–143) (50 µM) in 140 mM NaCl and 20 mM NaOAc, pH 3.7, was incubated at 25°C with... more <p>mPrP(107–143) (50 µM) in 140 mM NaCl and 20 mM NaOAc, pH 3.7, was incubated at 25°C without shaking alone (denoted by closed, half-filled and open squares for three independent measurements), in the presence of 30 µL of mPrP(23–230) seed (denoted by closed, half-filled and open circles for three independent measurements), or in the presence of the same amount of mPrP(23–230) seed digested by proteinase K (denoted by closed, half-filled and open up triangles for three independent measurements).</p

Skeletal muscle, 2016

Master transcription factor MyoD can initiate the entire myogenic gene expression program which d... more Master transcription factor MyoD can initiate the entire myogenic gene expression program which differentiates proliferating myoblasts into multinucleated myotubes. We previously demonstrated that histone methyltransferase KMT1A associates with and inhibits MyoD in proliferating myoblasts, and must be removed to allow differentiation to proceed. It is known that pro-myogenic signaling pathways such as PI3K/AKT and p38α MAPK play critical roles in enforcing associations between MyoD and transcriptional activators, while removing repressors. However, the mechanism which displaces KMT1A from MyoD, and the signals responsible, remain unknown. To investigate the role of p38α on MyoD-mediated differentiation, we utilized C2C12 myoblast cells as an in vitro model. p38α activity was either augmented via overexpression of a constitutively active upstream kinase or blocked via lentiviral delivery of a specific p38α shRNA or treatment with p38α/β inhibitor SB203580. Overexpression of KMT1A in ...

Acta neuropathologica, Jan 12, 2016

For proper mammalian brain development and functioning, the translation of many neuronal mRNAs ne... more For proper mammalian brain development and functioning, the translation of many neuronal mRNAs needs to be repressed without neuronal activity stimulations. We have discovered that the expression of a subclass of neuronal proteins essential for neurodevelopment and neuron plasticity is co-regulated at the translational level by TDP-43 and the Fragile X Syndrome protein FMRP. Using molecular, cellular and imaging approaches, we show that these two RNA-binding proteins (RBP) co-repress the translation initiation of Rac1, Map1b and GluR1 mRNAs, and consequently the hippocampal spinogenesis. The co-repression occurs through binding of TDP-43 to mRNA(s) at specific UG/GU sequences and recruitment of the inhibitory CYFIP1-FMRP complex by its glycine-rich domain. This novel regulatory scenario could be utilized to silence a significant portion of around 160 common target mRNAs of the two RBPs. The study establishes a functional/physical partnership between FMRP and TDP-43 that mechanistica...

Knockdown of p38α was monitored by western blot analysis of cell extracts from primary HsMB cells... more Knockdown of p38α was monitored by western blot analysis of cell extracts from primary HsMB cells expressing control scramble shRNA (Ctrl) or p38α shRNA, probed with antibodies for p38α, total p38, β-actin as loading control. Decreased levels of both p38α and total p38 were observed by p38α shRNA relative to Ctrl shRNA. (PPTX 47 kb)

Western blot analysis verified phosphorylated KMT1A by re-probing the membrane of the western blo... more Western blot analysis verified phosphorylated KMT1A by re-probing the membrane of the western blot results presented in Fig. 4a with a separate KMT1A antibody. (B) Western blot analysis of C2C12 cells expressing control scramble shRNA (Ctrl) or KMT1A shRNA via lentiviral delivery grown in GM or DM, probed with antibodies against KMT1A, and β-actin as loading control. Decreased levels of both under- and phosphorylated KMT1A were observed by KMT1A shRNA relative to Ctrl. (C) Flag-KMT1A overexpression was monitored by western blot analysis of cell extracts following it expression in 293A cells (293A-KMT1A-F) via lentiviral delivery. (D) In vitro kinase assays was performed for p38α activity using GST or GST-ATF2 as substrate by in vitro kinase assays in the presence and absence of SB. Commassie and autoradiography detected inputs GST/GST-ATF2 proteins and phosphorylated ATF2, respectively. (PPTX 99 kb)

(A) Ectopic Flag-KMT1A expression was determined by western blot analysis of C2-4RE-luc and C2-4R... more (A) Ectopic Flag-KMT1A expression was determined by western blot analysis of C2-4RE-luc and C2-4RE-Luc/KMT1A-F cells grown in GM, probed with antibodies to Flag-KMT1A, total KMT1A, MyoD, and β-actin as loading control. Flag-KMT1A expression was observed only in C2-4RE-luc/KMT1A-F cells. (B) Luciferase activity was monitored in C2-4RE-luc cells expressing vector control (Em), MKK6EE or MMK6DN grown in GM and values expressed after protein normalization as fold activation. Error bar, ±SEM (n = 3). MyoD-responsive reporter luciferase gene activation was observed by MKK6EE but not MKKDN in these cells. (PPTX 59 kb)

The principal event underlying the development of prion disease is the conversion of soluble cell... more The principal event underlying the development of prion disease is the conversion of soluble cellular prion protein (PrPC) into its disease-causing isoform, PrPSc. This conversion is associated with a marked change in secondary structure from predominantly a-helical to a high b-sheet content, ultimately leading to the formation of aggregates consisting of ordered fibrillar assemblies referred to as amyloid. In vitro, recombinant prion proteins and short prion peptides from various species have been shown to form amyloid under various conditions and it has been proposed that, theoretically, any protein and peptide could form amyloid under appropriate conditions. To identify the peptide segment involved in the amyloid core formed from recombinant full-length mouse prion protein mPrP(23–230), we carried out seed-induced amyloid formation from recombinant prion protein in the presence of seeds generated from the short prion peptides mPrP(107–143), mPrP(107–126), and mPrP(127–143). Our r...

Expression of HA-tagged MKK6EE and MKK6DN in C2C12 cells was verified by western blot analysis, p... more Expression of HA-tagged MKK6EE and MKK6DN in C2C12 cells was verified by western blot analysis, probed with antibodies against HA, and β-actin as loading control. (B) Control IgG or anti-phospho-p38 immunoprecipitates retrieved from extracts of indicated cells grown in GM or DM were subjected to in vitro kinase assays to monitor p38 activation using GST-ATF2 as substrate. Autoradiography and Commassie detected phosphorylated ATF2 and GST-ATF2 protein, respectively. (PPTX 75 kb)

Cell Reports

Highlights d TDP-43 cooperates with FMRP to modulate anterograde transport of dendritic mRNP d TD... more Highlights d TDP-43 cooperates with FMRP to modulate anterograde transport of dendritic mRNP d TDP-43 and Staufen1 co-regulate retrograde transport of mRNP in neuronal dendrites d TDP-43 regulates kinetics and dynamics of coupled dendritic transport-translation d TDP-43 regulates spine base-to-spine transport of Rac1 mRNA for local translation Authors

Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanisms

The DNA methylation program in vertebrates is an essential part of the epigenetic regulatory casc... more The DNA methylation program in vertebrates is an essential part of the epigenetic regulatory cascade of development, cell differentiation, and progression of diseases including cancer. While the DNA methyltransferases (DNMTs) are responsible for the in vivo conversion of cytosine (C) to methylated cytosine (5mC), demethylation of 5mC on cellular DNA could be accomplished by the combined action of the ten-eleven translocation (TET) enzymes and DNA repair. Surprisingly, the mammalian DNMTs also possess active DNA demethylation activity in vitro in a Ca2+- and redox conditions-dependent manner, although little is known about its molecular mechanisms and occurrence in a cellular context. In this study, we have used LC-MS/MS to track down the fate of the methyl group removed from 5mC on DNA by mouse DNMT3B in vitro and found that it becomes covalently linked to the DNA methylation catalytic cysteine of the enzyme. We also show that Ca2+ homeostasis-dependent but TET1/TET2/TET3/TDG-independent demethylation of methylated episomal DNA by mouse DNMT3A or DNMT3B can occur in transfected human HEK 293 and mouse embryonic stem (ES) cells. Based on these results, we present a tentative working model of Ca2+ and redox conditions-dependent active DNA demethylation by DNMTs. Our study substantiates the potential roles of the vertebrate DNMTs as double-edged swords in DNA methylation-demethylation during Ca2+-dependent physiological processes.

International Journal of Molecular Sciences

The intrinsic cellular heterogeneity and molecular complexity of the mammalian nervous system rel... more The intrinsic cellular heterogeneity and molecular complexity of the mammalian nervous system relies substantially on the dynamic nature and spatiotemporal patterning of gene expression. These features of gene expression are achieved in part through mechanisms involving various epigenetic processes such as DNA methylation, post-translational histone modifications, and non-coding RNA activity, amongst others. In concert, another regulatory layer by which RNA bases and sugar residues are chemically modified enhances neuronal transcriptome complexity. Similar RNA modifications in other systems collectively constitute the cellular epitranscriptome that integrates and impacts various physiological processes. The epitranscriptome is dynamic and is reshaped constantly to regulate vital processes such as development, differentiation and stress responses. Perturbations of the epitranscriptome can lead to various pathogenic conditions, including cancer, cardiovascular abnormalities and neurol...