Amita Limaye

• National Centre for Cell Science

People in developing countries have faced multigenerational undernutrition and are currently undergoing major lifestyle changes, contributing to an epidemic of metabolic diseases, though the underlying mechanisms remain unclear. Using a Wistar rat model of undernutrition over 50 generations, we show that Undernourished rats exhibit low birth-weight...

Special AT-rich binding protein 1 (SATB1) is a global chromatin organizer and a transcription factor regulated by interleukin-4 (IL-4) during the early T helper 2 (Th2) cell differentiation. Here we show that SATB1 controls multiple IL-4 target genes involved in human Th cell polarization or function. Among the genes regulated by SATB1 is that enco...

The chromatin organizer SATB1 regulates distant genes by selectively tethering matrix attachment regions (MARs) to the nuclear matrix. Post-translational modifications (PTMs) are important regulators of functional activities of proteins. Recently, a phosphorylation-dependent molecular switch that provided insights into the molecular mechanism of tr...

Qualitative analysis of proteins used for in vitro pulldowns. Coomassie stained gels depicting the purity of various GST-β-catenin (lanes 3–5, 9, 10) and SATB1 (lanes 13, 14) truncations used in GST pulldown experiments. His-SATB1 corresponds to 6× histidine-tagged full-length SATB1, CD+HD is the C-terminal half of SATB1 containing the DNA-binding...

Common targets of SATB1 and β-catenin. Expression profiling of cells in which β-catenin was overexpressed, knocked out, or mutated available on NCBI public domain GEO datasets GSE1473 [60] and GSE1579 [61] were analyzed against expression profiling of SATB1 overexpression series GSE4317 [21] as described below. Analysis of multiple arrays generated...

SATB1 and β-catenin interact in vivo. (A) Coimmunoprecipitation reactions was performed by separately incubating anti-β-catenin and anti-SATB1 antibodies with aliquots of Jurkat nuclear extract as described in Materials and Methods followed by Western blot (WB) using anti-SATB1 and anti-β-catenin, respectively. IgH, immunoglobulin heavy chain. (B)...

β-catenin activates SATB1-mediated transcription. The functional consequence of overexpression of SATB1 and constitutively active mutant β-catenin (T41A) on MAR-linked reporter assay was monitored as described in Materials and Methods. The IgH-MAR-luciferase construct consists of seven copies of the 25 bp AT-rich core of the IgH-MAR [62] that is a...

Analysis of SBSs in regulatory regions of multiple genes. EMSA analysis was performed using 32P-labeled upstream regulatory regions from CHUK, PPM1A, c-Myc genes, and the major breakpoint region of BCL2. Purified GST was used as a control for the GST:SATB1 fusion protein. The multiple regions tested from the 1 kb upstream region of CHUK and PPM1A a...

SATB1 and β-catenin occupy the SBSs in IL-2 and c-Myc promoters in vivo. (A) ChIP-PCR analysis in Jurkat cells showing the binding of SATB1 and β-catenin on IL-2 promoter in presence (+) and absence (−) of LiCl. ChIP analysis was performed as described in Materials and Methods. LiCl treatment resulted in increased occupancy of SATB1 at IL-2 promote...

Confirmation of polarization of CD4+ T cells to TH1 and TH2 subsets. Quantitation of marker cytokines in culture supernatants harvested from TH cells grown for 72 h was performed simultaneously using a multiplex bead array reader as described in Materials and Methods. As expected, TH1 cells produced IFN-γ and IL-12 whereas TH2 cells produced IL-4,...

SATB1 does not bind to the TCF consensus in vitro. In vitro binding analysis of TCF consensus binding site using recombinant SATB1 was performed by EMSA as described in Materials and Methods. The various panels depict EMSA using 32P-labeled wild-type (lanes 1–5) and mutant TCF (lanes 6–10) consensus sequences. 32P-labeled IgH-MAR was used as a posi...

Analysis of c-Maf expression in differentiating TH cells. Naïve CD4+ T cells were transfected with duplex siSATB1, siβ-catenin, or SATB1 overexpression plasmid DNA and differentiated ex vivo as described in Materials and Methods. As the control, duplex scrambled RNA (Scr) was transfected. Upon polarization for 72 h, total RNA was isolated and GATA-...

Analysis of upstream regulatory regions of GATA-3. In vitro binding analysis of GATA-3 promoter using recombinant SATB1 was performed by EMSA as described in Materials and Methods. The binding of SATB1 across the 2 kb (+1 to −2,000 bp) upstream regulatory region of GATA-3 was monitored by using 32P-labeled fragments of this region. We focused on a...

List of genes and gene IDs. The genes studied here or mentioned in the text are tabulated along with their respective gene IDs. (0.05 MB DOC)

SATB1 and β-catenin colocalize in the nuclei of mouse thymocytes. Triple stained immunofluorescence image (Alexa 488, Alexa 546, and DAPI). Thymocyte nuclei were counterstained with DAPI to mark the DAPI intense and less intense areas in the nuclei. The movie depicts a 3-D stack of images from top to bottom in the same thymocytes as in Figure 1A. T...

Human thymocytes were isolated from neonatal thymii as described in Materials and Methods and co-cultured with the OP9 or OP9-DL1 cells as described [63]. In brief, thymocytes were cultured on monolayers of OP9 or OP9-DL1 cells for 48 h in RPMI 1640 medium (Invitrogen) supplemented with 10% FCS, along with Wnt3a (10 µg/ml) (Wnt agonist) or DKK1 (10...

IL-4 is downregulated upon siRNA-mediated silencing of SATB1 in differentiating TH2 cells. Human CD4+ T cells were transfected with Scrambled (Scr) and si SATB1 synthetic duplex RNAs and then polarized to TH2 as described in Materials and Methods. After 72 h the cells were harvested and used for monitoring the expression of IL-4. (A) IL-4 gene expr...

β-catenin forms a characteristic “cage-like” substructure in the nuclei of mouse thymocytes. Single stained immunofluorescence image (Alexa 488). The movie depicts a 3-D stack of images from top to bottom in the same thymocytes as in Figure 1A. Images were captured using the Apotome module (Carl Zeiss) as described above. β-catenin staining pattern...

In vertebrates, the conserved Wnt signalling cascade promotes the stabilization and nuclear accumulation of beta-catenin, which then associates with the lymphoid enhancer factor/T cell factor proteins (LEF/TCFs) to activate target genes. Wnt/beta -catenin signalling is essential for T cell development and differentiation. Here we show that special...

Naïve CD4+ Th progenitors (Thp) differentiate to functionally distinct effector T cell subsets including Th1, Th2 and Th17 cells which while being responsible for specific immune functions have also been implicated in pathological responses, such as autoimmunity, asthma and allergy. Special AT-rich binding protein 1 (SATB1) is a global chromatin or...

Special AT-rich binding protein 1 (SATB1) acts as a global regulator of gene expression by recruiting various corepressor or coactivator complexes, thereby establishing a unique chromatin structure at its genomic targets in a context-dependent manner. Although SATB1 acts predominantly as a repressor via recruitment of histone deacetylase 1 (HDAC1)...

Chlorophenols and their derivatives are a major component of environmental pollutants that are potential immunotoxicants. Deaminase assay performed on peripheral blood mononuclear cells (PBMCs) exposed to chlorophenolic compounds and its derivatives demonstrated a decreased proliferation rate and cell death. Chlorophenolic exposure also led to impa...

SATB1 regulates gene expression by acting as a "docking site" for several chromatin remodeling enzymes and also by recruiting corepressors (HDACs) or coactivators (HATs) directly to promoters. However, how these contrasting effectors act at the level of SATB1 is not clear. We show here that phosphorylation by PKC acts as a switch to determine wheth...