Biology subjects

Anakk, S.

Publications and source records attributed to Anakk, S..

12 recordsLinked to original sources

bioRxiv2025

Constitutive Androstane Receptor induces Ribonucleotide Reductase-M2 expression and maintains hepatocyte ploidy in mice

The nuclear receptor Constitutive Androstane Receptor (CAR/NR1i3) is known for regulating various liver functions, including detoxification, nutrient metabolism, and hepatocyte proliferation. While CAR activation has been previously linked to higher ploidy, the underlying mechanisms are not fully known. Here, we uncover a basal role for CAR in maintaining hepatocyte ploidy, such that CAR deletion increases the number of diploid (2c) hepatocytes with a concomitant reduction in tetraploid (4c) hepatocytes. We demonstrate that CAR controls the de novo dNTP synthesis by directly transactivating the Ribonucleotide Reductase-M2 (RRM2) gene, which encodes the rate-limiting catalytic subunit of the enzyme, ribonucleotide reductase. Further, we find that the ligand-dependent CAR activation is sufficient to induce several genes involved in the de novo dNTP synthesis pathways, resulting in higher hepatic dATP and dTTP levels within the liver. Importantly, overexpressing RRM2 levels in CAR knockouts led to increased DNA synthesis and tetraploid (4c) hepatocytes compared to the control mice. Importantly, we demonstrate that CAR-mediated DNA synthesis in liver cells is dependent on catalytically active RRM2 function. Taken together, these findings reveal that the CAR-mediated RRM2 regulation contributes towards DNA synthesis and thereby maintains hepatocyte ploidy. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=68 SRC="FIGDIR/small/651109v2_ufig1.gif" ALT="Figure 1"> View larger version (29K): org.highwire.dtl.DTLVardef@1750110org.highwire.dtl.DTLVardef@98f579org.highwire.dtl.DTLVardef@1c0e87org.highwire.dtl.DTLVardef@1fdfda5_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

bioRxiv2024

Portal bile acids and microbiota along the murine intestinal tract exhibit sex differences in physiology

Microbes in the intestine transform bile acids during transit, altering their functional and signaling capacities before absorption into the portal vein. Sex differences in the gut microbiota have been noted, but their consequence on bile acid composition is unclear. Here, we investigated the composition and imputed functional potential of microbes in the small and large intestine together with portal and systemic bile acids. Female and male mice exhibit distinct microbial diversity throughout the length of the intestine leading to dimorphism in genes related to bile acid transformation. Subsequently, we found that the total portal bile acid concentrations were doubled in female mice compared to males. Conversely, oxo-bile acids that are rare in systemic circulation represented almost 30 percent of the portal pool in the male mice. Oxo- and deconjugated-bile acids were absent in germ-free mice consistent with microbe-mediated bile acid transformation. More importantly, gnotobiotic mice do not show sex differences in portal bile acids. Taken together, we demonstrate that sex differences in gut microbiota cause dimorphism in bile acid levels within the enterohepatic loop. HighlightsO_LIMurine microbial diversity exhibits sex-specific patterns along the small intestinal tract and colon. C_LIO_LIDistinct gut microbial profiles confer differential abundance of secondary bile acid metabolism genes in male and female mice. C_LIO_LIPortal bile acid compositions mirror sex differences in microbe-mediated bile acid processing that are lost in germ free conditions. C_LI

physiology↗

bioRxiv2021

Liver-specific Deletion Of Small Heterodimer Partner Alters Enterohepatic Bile Acid Levels And Promotes Bile Acid-Mediated Proliferation In Male Mice.

Small heterodimer partner (Shp) regulates several metabolic processes, including bile acid levels, but lacks the conserved DNA binding domain. Phylogenetic analysis revealed conserved genetic evolution of Shp, Fxr, Cyp7a1 and Cyp27a1, underscoring the importance of these molecules in maintaining bile acid homeostasis. Shp, although primarily studied as a downstream target of Farnesoid X Receptor (Fxr), has a distinct hepatic role that is poorly understood. Here we report that liver-specific Shp knockout (LShpKO) mice have impaired negative feedback of Cyp7a1 and Cyp8b1 upon bile acid challenge and demonstrate that a single copy of the Shp gene is sufficient to maintain this response. LShpKO mice also exhibit elevated total bile acid pool with higher bile acid fraction in the intestine mimicking the 1% cholic acid (CA) fed control mice. Agonistic activation of Fxr (GW4064) in the LShpKO did not alter the elevated basal expression of Cyp8b1 but lowered Cyp7a1 expression. We found that deletion of Shp led to an enrichment of distinct motifs and pathways associated with circadian rhythm, amino and carboxylic acid metabolism, copper ion transport, and DNA synthesis. LShpKO livers displayed a higher basal proliferation that was exacerbated specifically with bile acid challenge but not with another liver mitogen, TCPOBOP (TC). Overall, our data indicate that hepatic SHP uniquely regulates certain proliferative and metabolic cues.

physiology↗

bioRxiv2021

Lipid droplet structural remodeling in adipose tissue upon caloric excess

Excess calories are stored as triacylglycerols (TAG) and cholesteryl esters (CE) in lipid droplets (LD), and during obesity, LD expansion occurs. X-ray scattering of adipose tissue uncovered that LDs comprise two TAG packing domains: a disordered core and a multilamellar shell. The number of TAG layers increases upon diet-induced obesity and is adipose depot-specific. Further, collagen was highly oriented in brown but randomly dispersed in white fat. We discovered that the bodys surfactant, bile acids (BAs) stimulate remodeling of LD size. Deleting the BA receptor, Farnesoid X receptor (FXR) reduced a hydrophilic BA, {beta} muricholic acid ({beta}-MCA), and enlarged the adipocytes. BA composition is a critical determinant of overall hydrophobicity index and solubilization ability. Accordingly, we found that the obesogenic diet reduced a hydrophobic BA, chenodeoxycholic acid (CDCA). Taken together, these findings implicate that BAs, tissue niches, and diet influence LD structural remodeling. SummaryLipid droplets (LDs) pack triacylglycerols (TAGs) with altered dimensions and exhibit distinct collagen orientation between the white and brown fat depots and are remodeled by bile acids (BAs) such that deletion of BA-receptor, Farnesoid X receptor (FXR) results in adipocyte hypertrophy.

biophysics↗

bioRxiv2021

Splicing Factor SRSF1 Deficiency in the Liver Triggers NASH-like Pathology via R-Loop Induced DNA Damage and Cell Death

Regulation of RNA processing contributes profoundly to tissue development and physiology. Here, we report that serine-arginine-rich splicing factor 1 (SRSF1) is essential for hepatocyte function and survival. Although SRSF1 is mainly known for its many roles in mRNA metabolism, it is also crucial for maintaining genome stability. We show that acute liver damage in the setting of targeted SRSF1 deletion in mice is primarily mediated by the excessive formation of deleterious RNA-DNA hybrids (R-loops), which induce DNA damage. Combining hepatocyte-specific transcriptome, proteome, and RNA binding analyses, we demonstrate that widespread genotoxic stress following SRSF1 depletion results in global inhibition of mRNA transcription and protein synthesis, leading to impaired metabolism and trafficking of lipids. Lipid accumulation in SRSF1-deficient hepatocytes is followed by necroptotic cell death, inflammation, and fibrosis, resulting in NASH-like liver pathology. Importantly, SRSF1-depleted human liver cancer cells recapitulate this pathogenesis illustrating a conserved and fundamental role for SRSF1 in preserving genome integrity and tissue homeostasis. Thus, our study uncovers how accumulation of detrimental R-loops impedes hepatocellular gene expression, triggering metabolic derangements and liver failure.

cell biology↗

bioRxiv2020

Bile acid excess impairs thermogenic function in brown adipose tissue

Bile acids (BAs) not only facilitate fat digestion but also protect against obesity. Here, we show that a genetic mouse model for BA overload (Farnesoid X receptor; Small heterodimer double knockout (DKO)) exhibits mitochondrial dysfunction resulting in a thermogenic defect. By housing DKO mice at thermoneutrality, the poor mitochondrial function in brown fat protects them from diet-induced obesity. Compared to control, we find higher adipose BA levels with excess accumulation of taurocholic acid in the DKO mice. We report that the expression of genes responsible for BA de novo synthesis, conjugation and transporters and accumulation of BAs are present in both brown and white adipocytes. We determine that BA overload is sufficient to cause adipocyte mitochondrial dysfunction and induce the expression of cellular senescence genes in vitro. Taken together, we uncover that BA levels within the adipose tissue may modulate its overall function. HighlightsO_LIMouse model of BA overload exhibits adipose defects, which is partially restored by housing at thermoneutrality. C_LIO_LIBAs are present in detectable concentrations in both BAT and WAT. C_LIO_LIAdipocytes express genes responsible for de novo synthesis, conjugation and transport of BAs, and accumulate BAs. C_LIO_LIPathological accumulation of BAs impairs mitochondrial function leading to thermogenic defect. C_LI

physiology↗

bioRxiv2020

Deletion of intestinal SHP impairs short-term response to cholic acid challenge in mice.

Small heterodimer partner (SHP) is a crucial regulator of bile acid (BA) transport and synthesis; however, its intestine-specific role is not fully understood. Here, we report that Intestine-specific Shp knockout (IShpKO) mice have higher intestinal and hepatic BAs, but not serum BAs when challenged with an acute (5-day) 1% cholic acid (CA) diet. Consistent with this finding, BA synthetic genes Cyp7A1 and Cyp8b1 are not repressed to the same extent in IShpKO compared to control mice post-CA challenge. Loss of intestinal SHP did not alter Fxr mRNA but increased Asbt (BA ileal uptake transporter) and Ost (BA ileal efflux transporter) expression even under chow-fed conditions. Surprisingly, the acute CA diet in IShpKO did not elicit the expected induction of Fgf15 but was able to maintain the suppression of Asbt, and Ost/{beta} mRNA levels. At the protein level, ASBT was downregulated, while OST/{beta} expression was induced and maintained regardless of diet. Examination of ileal histology in IShpKO mice challenged with acute CA diet revealed reduced villus length and goblet cell numbers. However, no difference in goblet cell number, villus morphology, crypt depth, and the expression of BA regulator and transporter genes was seen between f/f Shp and IShpKO mice after chronic (14-day) CA diet suggesting an adaptive response. We found the upregulation of the Ppar-Ugt axis, which can reduce the BA burden and compensate for the ileal SHP function. Thus, our study reveals that ileal SHP expression contributes to both overall intestinal structure and BA homeostasis.

physiology↗

bioRxiv2020

Circulating bile acid levels direct sex-differences in liver cancer development

Hepatocellular carcinoma (HCC), the most common liver cancer, exhibits a higher incidence in males. Here, we report that mice lacking the bile acid regulators, Farnesoid X Receptor (FXR) and Small Heterodimer Partner (SHP), recapitulate the sex difference in liver cancer risk. Since few therapeutic options are available, we focused on understanding the intrinsic protection afforded to female livers. Transcriptomic analysis in control and FXR and SHP double knockout livers identified female-specific changes in metabolism, including amino acids, lipids, and steroids. We examined if the obtained transcriptomic signatures correlate with the survival outcomes for HCC patients to assess the translational potential of this murine HCC model. Gene signatures that are unique to the knockout females correspond with low-grade tumors and better survival. Ovariectomy blunts the metabolic changes in female livers and promotes tumorigenesis that, intriguingly, coincides with increases in serum bile acid (BA) levels. Despite similar genetics, we found higher serum BA concentrations in males, whereas female knockout mice excreted more BAs. Decreasing enterohepatic BA recirculation using cholestyramine, an FDA-approved resin, dramatically reduced the liver cancer burden in male mice. Overall, we reveal that sex-specific BA metabolism leading to lower circulating BA concentration protects female livers from developing cancer. Thus, targeting BA excretion may be a promising therapeutic strategy against HCC. SignificanceWe show that female-specific gene profiles identified in Fxr-/-, Shp-/- double knockout (DKO) mice correlate with better outcomes for HCC patients and uncover sex differences in circulation and excretion of bile acid. Overall, we demonstrate that increasing (ovariectomy or chemical injury) or decreasing (pharmacologically with FDA-approved resin) serum bile acids, not hepatic bile acids, promoted or alleviated liver cancer burden.

cancer biology↗

bioRxiv2020

Deletion and overexpression of the scaffolding protein IQGAP1 promotes HCC

IQ motif-containing GTPase-activating protein 1 (IQGAP1) is a ubiquitously expressed scaffolding protein that is overexpressed in a number of cancers, including liver cancer, and is associated with many pro-tumorigenic processes including cell proliferation, motility, and adhesion. IQGAP1 can integrate multiple signaling pathways and could be an effective anti-tumor target. Therefore, we examined the role for IQGAP1 in tumor initiation and promotion during liver carcinogenesis. Unexpectedly, we found that Iqgap1-/- mice had a higher tumor burden than Iqgap1+/+ and Iqgap1+/- mice following DEN-induced liver carcinogenesis. Iqgap1-/- tumors as well as knocking down IQGAP1 in hepatocellular carcinoma (HCC) cell lines resulted in increased MET activation and cellular proliferation. On the other hand, we uncovered IQGAP1 overexpression accelerates HCC development by YAP activation and subsequent NUAK2 expression. We demonstrate that increasing IQGAP1 expression in vivo does not alter {beta}-catenin or MET activation. Taken together, we identify that both loss and gain of function of IQGAP1 promotes HCC development by two separate mechanisms in the liver. These results demonstrate that adequate amount of IQGAP1 is necessary to maintain a quiescent status of liver.

cancer biology↗

bioRxiv2020

Scaffolding protein IQ motif containing GTPase activating protein 2 (IQGAP2) regulates glycogen metabolism

The liver is critical in maintaining metabolic homeostasis, regulating both anabolic and catabolic processes of fats, proteins, and carbohydrates. The IQ motif containing GTPase activating protein 2 (IQGAP2) is a member of the IQGAP family. Of the three homologous isoforms, the IQGAP2 scaffolding protein is predominantly found in the liver. To characterize its role in regulating metabolism, Iqgap2-/- female and male mice, and their WT controls, were fed ad libitum or fasted for 24 hours. Hepatic gene expression, protein levels, and the metabolic response were compared between WT and Iqgap2-/- mice, using RT-qPCR, western blot analysis, and histological stains. We found that loss of IQGAP2 alters the phosphorylation of active glycogen synthase kinase 3 (GSK3) expression, a known regulator of glycogen synthesis and lipogenesis. Consistent with this result, Iqgap2-/- female mice displayed depletion of periportal glycogen even in the fed state. We also observed the blunted expression of genes involved in glycogenesis and lipogenesis when IQGAP2 was deleted. Since GSK3 is known to regulate the activity of {beta}-catenin, we examined and found it to be reduced in Iqgap2-/- mice. Our findings demonstrate that IQGAP2 plays an important role in regulating glycogen synthesis.

molecular biology↗

bioRxiv2019

Constitutive Androstane Receptor contributes towards increased drug clearance in cholestasis

Understanding the alterations in drug metabolism in different liver diseases is crucial for appropriate therapeutic intervention. We performed high-throughput RNA sequencing on various liver injury models, including cholestasis, diet-induced steatosis, and regeneration. Comparative liver transcriptome analysis revealed overlapping and distinct gene profiles among different liver diseases. Particularly, cholestatic livers displayed robust induction of drug metabolizing genes. This upregulation is not a generic hepatic stress response, as it was suppressed or unchanged in other models of liver diseases. Consistently, drug metabolic gene profiles were induced in a subset of biliary atresia patients, but not in individuals with hepatitis B or C viral infection, and alcoholic hepatitis. Further analysis revealed this induction was specific to genes regulated by nuclear receptor CAR (Constitutive Androstane Receptor). To test this, we challenged cholestatic mice with a paralytic agent, zoxazolamine. Compared to controls, these mice displayed significantly reduced paralysis time, reflecting increased drug metabolism, and this effect was lost upon inhibition of CAR. Thus, CAR activation can alter therapeutic efficacy of certain drugs in a subset of cholestatic individuals.

physiology↗

bioRxiv2017

Deletion of murine IQGAP1 results in increased mTOR activation and blunted ketogenic response

IQ motif-containing GTPase Activating Protein 1 (IQGAP1) is a ubiquitously expressed scaffolding protein that integrates signaling from multiple cellular processes including motility, adhesion, and proliferation. Here, we show that IQGAP1 is induced in the liver upon fasting and can also regulate {beta}-oxidation and ketone body synthesis. Utilizing ketogenic diet and pharmacologic activation we identified that hepatic PPAR activity is compromised in Iqgap1-/-mice. Our data show that IQGAP1 interacts with the mechanistic target of rapamycin (mTOR) and IQGAP1 deletion results in enhanced mTOR complex 1 (mTORC1) activation. Conversely, ectopic expression of IQGAP1 in Iqgap1-/- mice was sufficient to suppress mTORC1 signaling. We also confirmed that modulation of mTORC1 signaling by IQGAP1 is cell autonomous. This increased mTORC1 activation impedes PPAR signaling since mTORC1 inhibition restored a subset of metabolic genes in Iqgap1-/- mice. Overall, we demonstrate a previously unidentified role for IQGAP1 as an important regulator of mTORC1 activity and long-term ketosis.

physiology↗