SUMMARY Commensal gut microflora and dietary fiber protect against colonic inflammation and colon cancer through unknown targets. Butyrate, a bacterial product from fermentation of dietary fiber in the colon, has been implicated in this process. GPR109A (encoded by Niacr1) is a receptor for butyrate in the colon. GPR109A is also a receptor for niacin, which is also produced by gut microbiota and suppresses intestinal inflammation. Here we showed that Gpr109a signaling promoted anti-inflammatory properties in colonic macrophages and dendritic cells and enabled them to induce differentiation of Treg cells and IL-10-producing T cells. Moreover, Gpr109a was essential for butyrate-mediated induction of IL-18 in colonic epithelium. Consequently, Niacr1−/− mice were susceptible to development of colonic inflammation and colon cancer. Niacin, a pharmacological Gpr109a agonist, suppressed colitis and colon cancer in a Gpr109a-dependent manner. Thus, Gpr10a has an essential role in mediating the beneficial effects of gut microbiota and dietary fiber in colon.

ATB 0,ϩ (amino acid transporter responsible for the activity of system B 0,ϩ ) was named "B 0,ϩ " to indicate its broad substrate selectivity (denoted by "B"), accepting neutral (denoted by "0") and cationic (denoted by "ϩ") amino acids as substrates (1, 2). Its transport function is coupled to a Na ϩ gradient, a Cl Ϫ gradient, and membrane potential. ATB 0,ϩ is identified as SLC6A14 according to the Human Genome Organization nomenclature. This transporter has potential for delivery of a wide variety of drugs and prodrugs into cells (3-7). The substrate selectivity of SLC6A14 is interesting (1, 2). It transports all essential amino acids. The only excluded amino acids are glutamate and aspartate, which are nonessential. It also transports glutamine (an important precursor for nucleotide synthesis) and arginine (an amino acid essential for tumor growth). However, the transporter is expressed only at low levels in normal tissues. Tumor cells have an increased requirement for essential amino acids as well as glutamine and arginine to support their rapid growth. Essential amino acids are obligatory for protein synthesis. Leucine, an essential amino acid, is also a potent activator of mTOR (mammalian target of rapamycin) (8). Certain tumor cells metabolize glutamine at a rate far exceeding the requirement for protein and nucleotide synthesis, a phenomenon known as "glutamine addiction" (9 -11). Glutamine metabolism through a set of biochemical reactions called "glutaminolysis" provides a carbon source for tumor cells, thereby sparing glucose-derived carbon for the synthesis of lipids and other essential biomolecules. Arginine is essential for several types of cancer due to lack of the arginine-synthesizing enzyme argininosuccinate synthetase (12, 13). On the basis of these findings, we hypothesized that the expression of SLC6A14 may be up-regulated in cancer to meet the increasing demand for all essential amino acids as well as glutamine and arginine. In support of this hypothesis, past studies from our laboratory have shown that SLC6A14 is up-regulated in colon cancer (14) and cervical cancer (15). The transporter is also up-regulated in breast cancer cell lines, but interestingly only in estrogen receptor (ER) 2 -positive cell lines (16). Furthermore, we showed that blockade of SLC6A14 in ER-positive breast cancer cells by treatment with a selective blocker (␣-methyl-DLtryptophan (␣-MT)) starved the cells of glutamine, arginine, and essential amino acids, decreased cell proliferation, and caused apoptotic cell death (16). In the present study, we investigated the expression of SLC6A14 in primary breast cancer * This work was supported, in whole or in part, by National Institutes of Health Grant CA152396. 1 To whom correspondence should be addressed. E-mail: vganapat@ georgiahealth.edu.2 The abbreviations used are: ER, estrogen receptor; ␣-MT, ␣-methyl-DL-tryptophan; CHOP, CCAAT/enhancer-binding protein homologous protein; 3-MA, 3-methyladenine.

The Ufm1 conjugation system is a novel ubiquitin-like modification system, consisting of Ufm1, Uba5 (E1), Ufc1 (E2) and poorly characterized E3 ligase(s). RCAD/Ufl1 (also known as KIAA0776, NLBP and Maxer) was reported to function as a Ufm1 E3 ligase in ufmylation (Ufm1-mediated conjugation) of DDRGK1 and ASC1 proteins. It has also been implicated in estrogen receptor signaling, unfolded protein response (UPR) and neurodegeneration, yet its physiological function remains completely unknown. In this study, we report that RCAD/Ufl1 is essential for embryonic development, hematopoietic stem cell (HSC) survival and erythroid differentiation. Both germ-line and somatic deletion of RCAD/Ufl1 impaired hematopoietic development, resulting in severe anemia, cytopenia and ultimately animal death. Depletion of RCAD/Ufl1 caused elevated endoplasmic reticulum stress and evoked UPR in bone marrow cells. In addition, loss of RCAD/Ufl1 blocked autophagic degradation, increased mitochondrial mass and reactive oxygen species, and led to DNA damage response, p53 activation and enhanced cell death of HSCs. Collectively, our study provides the first genetic evidence for the indispensable role of RCAD/Ufl1 in murine hematopoiesis and development. The finding of RCAD/Ufl1 as a key regulator of cellular stress response sheds a light into the role of a novel protein network including RCAD/Ufl1 and its associated proteins in regulating cellular homeostasis. Cell Death and Differentiation (2015) 22, 1922-1934 doi:10.1038/cdd.2015 .51; published online 8 May 2015The Ufm1 (Ubiquitin-fold modifier 1) conjugation system is a novel ubiquitin-like (Ubl) modification system that shares biochemical features with other Ubl systems.1 Ufm1 modifies its target proteins through a biochemical pathway catalyzed by specific E1 (Uba5), E2 (Ufc1) and E3 enzyme(s) even though the identities of E3 ligases remain mostly elusive. Genetic study from Uba5 knockout (KO) mice has shown that Uba5 is indispensable for embryonic erythropoiesis, highlighting the pivotal role of this novel Ubl system in animal development. 2Yet its role in adult erythropoiesis and other developmental processes is largely unexplored and the underlying molecular mechanism remains poorly understood.Regulator of C53 and DDRGK1 (also known as KIAA0776, Ufl1, NLBP and Maxer, referred to as RCAD hereafter) has recently been identified by independent studies as an important regulator of several signaling pathways, including protein ufmylation, NF-κB signaling and unfolded protein response (UPR).3-9 Endogenous RCAD forms a complex with two proteins: C53 (also known as LZAP and Cdk5rap3) 5,6,10 and DDRGK1 (also designated as C20orf116, Dashurin and UFBP1), 3,6,7,11 and regulates the stability of its binding partners. 5,6 Intriguingly, Tatsumi et al. 3 found that Ufl1 (same as RCAD) promoted ufmylation of DDRGK1, suggesting that RCAD may function as an E3 ligase for ufmylation of DDRGK1. In line with its role in ufmylation, knockdown of endogenous RCAD resulted in attenuated ufmylation of endogen...

Composition of the gut microbiota has profound effects on intestinal carcinogenesis. Diet and host genetics play critical roles in shaping the composition of gut microbiota. Whether diet and host genes interact with each other to bring specific changes in gut microbiota that affect intestinal carcinogenesis is unknown. Ability of dietary fibre to specifically increase beneficial gut microbiota at the expense of pathogenic bacteria in vivo via unknown mechanism is an important process that suppresses intestinal inflammation and carcinogenesis. Free fatty acid receptor 2 (FFAR2 or GPR43) is a receptor for short-chain fatty acids (acetate, propionate and butyrate), metabolites of dietary fibre fermentation by gut microbiota. Here, we show FFAR2 is down modulated in human colon cancers than matched adjacent healthy tissue. Consistent with this, Ffar2−/− mice are hypersusceptible to development of intestinal carcinogenesis. Dietary fibre suppressed colon carcinogenesis in an Ffar2-dependent manner. Ffar2 played an essential role in dietary fibre-mediated promotion of beneficial gut microbiota, Bifidobacterium species (spp) and suppression of Helicobacter hepaticus and Prevotellaceae. Moreover, numbers of Bifidobacterium is reduced, whereas those of Prevotellaceae are increased in human colon cancers than matched adjacent normal tissue. Administration of Bifidobacterium mitigated intestinal inflammation and carcinogenesis in Ffar2−/− mice. Taken together, these findings suggest that interplay between dietary fibre and Ffar2 play a key role in promoting healthy composition of gut microbiota that stimulates intestinal health.