Does Propionate affect the Expression of NRF-2 - Essay Example

Does Propionate affect the Expression of NRF-2? The carbohydrates that are not digested in the ileum are passed on tothe colon where they go through fermentation with the aid of bacteria. The resulting products are called short-chain fatty acids (SCFAs). Most of these SCFAs have not more than six carbon atoms. The common SCFAs are propionate, butyrate, and acetate. Once they are absorbed into the colonic epithelial cells, they undergo metabolism. The remaining SCFAs move on to the liver where metabolism also occurs in the hepatocytes. Their effects and functions are diverse and the subject of numerous studies. It is through their effects that the review looks at the nuclear response factor-2 (NRF-2). What effect do they have on the expression of NRF-2? Keywords: Short fatty chain acids (SCFAs), GA-Binding protein (GABP) or nuclear response factor (NRF-2), non-digestible carbohydrate (NDC) Does Propionate affect the Expression of NRF-2? Introduction According to Reichdart, et al. (2014), propionate is secreted in the human colon through microbial fermentation. Propionate is a short-chain fatty acid (SCFA), which along with two other, acetate and butyrate, are referred to as the common short fatty acids. However, what are short fatty acids? Short-chain fatty acids are fatty acids that have less than eight carbons. The number of carbons in fatty acids affect their physical functions along with their chemical characteristics (Stewart, 2008). In the case of propionate, it has three carbon atoms. Propionate makes up twenty percent of the total SCFAs, while the other two constitute the rest (den Besten, et al., 2013). As propionate has been described, the question that arises is what is NRF-2? Nuclear respiratory factor 2 (NRF-2) is also known as GA binding protein. This is according to Priya, Johar, and Wong-Riley (2013) who state that NRF-2 is a factor of transcription found in the family known as the E-26 transformation-specific. As a protein, it has a functional component made up of a β and an α sub-unit. The two sub-units can form a heterotetramer or a heterodimer. As indicated, the area involved with the triggering of transcription is located in the β sub-unit. On the other hand, the α sub-unit comprises the field, ETS DNA binding field, which binds to either the TTCC or the GGAA motif. As a transcription factor, NRF-2 regulates various cellular processes like the synthesis of proteins and the cell cycle. The protein is located in specific neurones that function to control translocation of the nuclear and transcription of it’s α and β sub-units (Bruni, et al., 2010). The main question that the paper now seeks to answer is whether the SCFA propionate, affects the expression of NRF-2. Expression and Cloning of NRF-2 Gugneja, et al. (1995) successfully cloned NRF-2 with both β and α subunits. The researchers utilised a PCR that had HeLA cells. With the help of degenerate primers developed from the existing sequences of amino acids obtained of refined GABP proteins, they were able to clone the GA-Binding Protein 2. However, over the years, it had always been thought the cloned NRF-2 were either of the two variants, α and β. Nevertheless, on further research by various studies such as one conducted by Sementchenko and Watson in the year 2000, it was evaluated that the four splice variants were all under the gene GABPB. Watanabe, et al. (1993) were able to clone HeLA cell cDNAs. Accrording to the authors, the cloned cDNAs were all involved in the encoding of several E4TF1 subunits. The subunits encoded by the cloned HeLA cDNAs included the GA-Binding Protein Beta-1, GA-Binding Protein Alpha, and GA-Binding Protein-2. The subunits GABPB-2 and GABPB-1 are said to comprise of 347 and 383 amino acids respectively. In addition, they are homologous in nature. It was found out that the N-terminal amino acids of both subunits were similar. These N-terminal amino acids comprised of four Notch motifs that repeat in tandem. The authors noted that the N-terminal amino acids were 332 in number. The only contrast that was evident was at the C termini, which was the only place that the two subunits varied. A major characteristic of the three E4FT1 proteins was that they expressed great homolgy with the related mice proteins that bind to GABP. The Role of NRF-2 NRF-2 has been found to associate with the subunit, GABPA, which binds to DNA. The importance of stating this is to note that NRF-2 does not bind to DNA directly. From research, it has been proved that there is activity that involves the binding of DNA in NRF-2 α subunit. In order for transcription to take place, either of the two NRF-2 sub-units is required. However, it is worthwhile to note that the subunit is not active when it comes to transcription. On further research, it has been discovered that all the variants of NRF-2 have similar capabilities when it comes to the triggering of transcription. It is especially so when they are linked to a domain that binds DNA in transfected cells. As Anuranjani and Bala, (2014) indicate, the variant form of NRF-2 that has the C-terminal leucine zipper motif is capable of homodimerising and coming up with a hetertetratmeric complex that is active for transcription. However, the NRF-2 α subunit is a variant that does not have the zipper motif. The subunit has the ability to form heterodimers. In spite of this capability, it is not able to come up with heterotramers that are active for transcription. Scarpulla, (2008) notes that the isorforms of NRF-2, which are the alpha and the beta, all function to steady GABPA’s binding affinity to DNA. The isoforms do this by conveying a decreased rate of dissociation. Through evaluation techniques such as Western and Northern blot, it was found that both NRF-2 subunits are expressed similarly. However, their expression occurs at varied ratios in some cells. Both of the NRF-2 subunits bind GABPA in vitro through a competitive mode. These variations in ratios between the two NRF-2 subunits has been said to influence the transcription from promoters that were triggered by GABP. Propionate Production in the Gut Research has associated non-digestible carbohydrates (NDC) to be the source of short-chain fatty acids. Hence, propionate been one of the major three SCFAs is not an exception. The NDCs are said to be non-digestible because they are not digested in the ileum where digestion takes place. However, they undergo fermentation in the colon with the help of bacteria that are found in the colon, which is a portion of the large intestine. These carbohydrates are first converted into pyruvate. Hence, pyruvate is the dominant metabolite that is used in the fermentation of carbohydrates. The focus been on propionate, one of the fermentation products, it is vital to note that the main bacteria species, which synthesises it, is the Bacteroides. However, propionate production is not limited to this species as two other bacteria species found in the colon have been associated with its synthesis. These species include the Veillonella and Propionibacterium. Several metabolic pathways are utilised in the fermentation of carbohydrates in the colon. They may include glycolysis, pentose phosphate pathway, or the oxidative decarboxylation of pyruvate although these are not the only metabolic pathways. The metabolic pathways that are utilised by bacteria to synthesise propionate are the acrylate pathway and the succinate pathway. The Bacteroides species has been known to make use of succinate pathway. Through this pathway, there is decarboxylation of succinate to produce propionate. On the other hand, propionate is also produced from lactate through the acrylate pathway. It is especially so for foods that have great quantities of oligosaccharides that cannot be digested (Rambaud, et al., 2006). Propionate Metabolism in the cells Once propionate is produced in the colon, it is readily absorbed like all other SCFAs. As Wong, et al. (2006) assert, the liver takes up most of the propionate. For propionate to be absorbed by liver hepatocytes, it must be taken up first by the epithelial cells. Studies involving the rumen epithelium show that before it is passed on to the hepatica portal circulation, propionate could go through metabolism in the epithelium. The metabolism in the epithelial cells may consist of changing it into an intermediate or oxidation. In spite of the knowledge that there could be metabolism of propionate in the epithelial cells, how it occurs has not yet been fully determined. It is the reason why some research studies demonstrate that propionate is mainly converted to lactate along with total oxidation to carbon (IV) oxide, production of pyruvate and its subsequent conversion to alanine. The most influencing factor in propionate metabolism in the epithelial cells is its concentration. Nevertheless, the SCFA butyrate has been seen to hinder the activation of propionate to propionyl-Coenzyme A. As a result, metabolism of propionate in epithelial cells is reduced. The hindrance leads to more release of propionate to hepatic portal circulation for transfer to hepatocytes (Burrin & Mersmann, 2005). The succinate pathway helps in the metabolism of propionate. In this pathway, propionate is initially changed to propionyl-Coenzyme A. The change is achieved through by reacting it with succinyl-Coenzyme A. After been changed, propionyl-Coenzyme A is reacted with oxaloacetate. The reaction yields methylmalonyl-Coenzyme A and pyruvate. According to Belaich, et al. (1990), methylmalonyl-Coenzyme A goes through an intricate intra-molecular reaction whereby it is rearranged and changed through a reaction involving mutase to produce succinyl-Coenzyme A. Succinyl-Coenzyme A is changed to succinate. In a study undertaken by Ali and Jois in 1997, it was demonstrated that propionate is metabolised in the liver in what is known as gluconeogenesis. The study found that glucagon influences highly on the metabolism of propionate in liver cells. When they treated the caudal lobes with glucagon, the uptake of propionate into hepatocytes increased by over fifty percent. The gluconeogenesis rate was also raised by more than thirty percent. The researchers calculated that out of the total propionate taken up by the hepatocytes; around sixty-three percent was produced as glucose. It was also found out that butyrate, another SCFA, was involved in the inhibition of propionate in the cells of the liver. Armentano (1992) is in support of the above findings as he notes that, the net uptake of propionate is a signal of the overall metabolism. The researcher notes that the hepatocytes are able to metabolise propionate. Nevertheless, an increase in diet containing NDCs results in the conversion of propionate into fat. It is changed into fat as a reaction to its intensified absorption. Overall, Wilkemeyer, et al. (1992) note that propionate metabolism comprises a sequence of enzymes located in the mitochondrial matrix like propionyl-Coenzyme A carboxylase and methylmalonyl-Coenzyme A mutase. In addition, in support of the above findings, Berry and Edwards (2000) assert that β-oxdiation is needed to back up metabolism of propionate in the liver cells to produce glucose. The authors smartly state that the metabolism of butyrate released intermediates that highly restricted production of glucose from propionate. Hence, they conclude that, the metabolism of propionate serves the purpose of availing energy for its change to glucose. Propionate and Cancer Cells From various studies that have been conducted on SCFAs, it has been noted that they are involved in the control of the expression of various genes. It is through this regulation that SCFAs have been linked to cancer. For example, one of the most examined SCFA is butyrate. The analysis undertaken shows that sodium butyrate controls various genes. The regulation wields an influence on the development of cancer of the colon. How does it do this? The compound sodium butyrate causes the hindrance of colon cancer cells growth. It does so through the enhancement of hyper-acetylation of histone. It also accomplishes the inhibition by introduction of the inhibitor that hinders the cycle of the cancer cells. The influence of butyrate, an SCFA, is just but one example. With the focus of the paper been on propionate, it has been suggested that propionate also hinders the growth of cancer cells. However, it does so to a reduced degree than butyrate. To examine the association between cancer and the SCFA propionate, Adom and Nie, (2013) treated colon cancer cells termed as HCT116 with propionate. On treatment, a decrease in the state of phosphorylation that was affected by time was observed. The reduced state of phosphorylation was detected at Ser2481. However, there was no alteration in the amount of the mammalian target of rapamycin (mTOR). The study also discovered that there was a decrease in phosphorylation of phospho-p70 S6 Kinase (p70S6K) at Thr389. What did they infer from these observations? The researchers inferred that the down-regulation of the signalling pathway of mTOR is a means that propionate utilises to cause cell death. The suggestion that the study supposed was that propionate has to cause the signalling of mTOR through the hindrance of the phosphatidylinositide 3-kinaseI3/Akt (PI3/Akt) pathway. However, propionate treatment enhanced the AMP-activated protein kinase (AMPK) leading to a reduction in the level of the energy producing molecule ATP in the cancer cells. It was achieved through the destruction of the potential of the mitochondrial membrane. As a result, when the mitochondria were stained, the intensity in the stain was little. It also resulted in an enhanced co-localisation between the punctuates and the mitochondria. One of the mitochondrial markers, COXIV, was decreased and targeted by auto-lysosomes as faulty mitochondria. The mitochondrial markers greatly co-localised with the protein-p62 that binds to ubiquitin. Thus, the study concluded that propionate activates mitophagy that was said to aim at the mitochondria that had a membrane potential, which was depolarised. The metabolism of lipids in the cancer cells was variated by propionate treatment. It was seen that the enzyme that regulates the production of long chain fatty acids, fatty acid synthase, was decreased. The researchers went a step further and examined the impact of propionate on biogenesis in the mitochondria. What did they find out? They found out that there was a boost in the mRNA quantity of the nuclear gene mitochondrial transcription factor A and factor B (Adom and Nie, 2013). Of importance to this paper was one crucial finding. The essential finding involved the NRF-2. The study specifically noted of the findings regarding certain transcription factors associated to the biogenesis of mitochondria and the bio-production of heme. In the research, it is specified that these transcription factors are NRF-2 and NRF-1 along with polymerase gamma. It is asserted that the expression of these transcription factors was enhanced at the mRNA level in the HCT116 cancer cells. The enhancement was after the treatment with propionate. In support of these findings was the research carried out by Tang, et al. (2011). In the study, the researchers report that propionate caused autophagy. They state that the autophagy was an indication of the therapeutic efficiency of propionate and SCFAs in general towards cancer of the colon. In agreement with the research conducted by Adon and Nie (2013), the study opines that the expression of NRF-2 and 1 at the mRNA level was increased. The study goes ahead to add that, the stimulation in the expression of NRF-2 was observed within 12 hours of propionate treatment. However, they emphatically note that the enhancement decreased after long-term treatment with propionate. Conclusion In conclusion, it can be said that propionate affects the expression of the nuclear response factor-2 by stimulating it at the mRNA level. The studies that have been discussed avail evidence for this stimulation through the treatment of cancer cells with propionate. The paper’s specific outlook on the effect of propionate on cancer cells and the NRF-2 is an indication of the overall effect of SCFAs. Even though their production from NDCs and the impact they have on each other’s functioning differ, the wholesome impact of SCFAs can be derived from that of propionate. It is worthwhile to note that not much research has been done on propionate as compared to that of butyrate. Hence, there is room for the analysis of the effects of propionate, not only in the expression of NRF-2, but also in normal and cancerous cells. References Adom, D. & Nie, D., 2013. Regulation of Autophagy by Short Chain Fatty Acids in Colon Cancer Cells. In: Y. Bailly, ed. Immunology and Microbiology. s.l.:INTECH Open Access Publisher, pp. 235-247. Ali, A. M. & Jois, M., 1997. Uptake abd metabolism of propionate in the liver isolated from sheep treated with glucagon. British Journal of Nutrition, Volume 77, pp. 783-793. Anuranjani & Bala, M., 2014. Concerted action of Nrf2-ARE pathway, MRN complex, HMGB1 and inflammatory cytokines - Implication in modification of radiation damage. Redox Biology, Volume 2, pp. 832-846. Armentano, L. E., 1992. Ruminant Hepatic Metabolism of Volatile Fatty Acids, Lactate and Pyruvate. The Journal of Nutrition, 122(3), pp. 838-842. Belaich, J.-P., Bruschi, M. & Garcia, J.-L. eds., 1990. Microbiology and Biochemistry of Strict Anaerobes Involved in Interspecies Hydrogen Transfer. New York: Springer Science & Business Media. Berry, M. N. & Edwards, A. M. eds., 2000. The Hepatocyte Review. Dordrecht: Kluwer Academic Publishers. Bruni, F. et al., 2010. Nuclear Respiratory Factor 2 Induces the Expression of Many but Not All Human Proteins Acting in Mitochondrial DNA Transcription and Replication*An external file that holds a picture, illustration, etc. Object name is sbox.jpg. Journal of Biological Chemistry, 285(6), pp. 3939-3948. Burrin, D. G. & Mersmann, H. J. eds., 2005. Biology of Metabolism in Growing Animals. Philadelphia: Elsevier Health Sciences. den Besten, G. et al., 2013. The role of short-chain fatty acids in the interplay between diet, gt microbiota and host energy metabolism. Journal of Lipid Research, 54(9), pp. 2325-2340. Gugneja, S., Virbasius, J. V. & Scarpulla, R. C., 1995. Four structurally distinct, non-DNA-binding subunits of human nuclear respiratory factor 2 share a conserved transcriptional activation domain.. Mollecular Cellular Biology, Volume 15, pp. 102-111. Priya, A., Johar, K. & Wong-Riley, M., 2013. Nuclear respiratory factor 2 regulates the expression of the same NMDA receptor subunit genes as NRF-1: Both factors act by a concurrent and parallel mechanism to couple energy metabolism and synaptic transmission. Biochimica et Biophysica Acta (BBA)- Molecular Cell Research, 1833(1), pp. 48-58. Rambaud, J.-C., Buts, J.-P., Corthier, G. & Flourie, B. eds., 2006. Gut Microflora: Digestive Physiology and Pathology. Surrey: John Libbey Eurotext. Reichdart, N. et al., 2014. Phylogenetic distribution of three pathways for propionate production within the human gut microbiota. The ISME Journal, Volume 8, pp. 1323-1335. Scarpulla, R. C., 2008. Transcriptional Paradigms in Mammalian Mitochondrial Biogenesis and Function. Psychological Reviews, 88(2), pp. 611-638. Sementchenko, V. I. & Watson, D. K., 2000. Ets target genes: past, present and future. Oncogene, 19(55), pp. 6533-6548. Stewart, M. L., 2008. In Vitro and in Vivo Studies of Short-chain Fatty Acid Production from Novel Resistant Starches as a Marker of Fermentation. Ann Arbor: ProQuest. Tang, Y., Chen, Y., Jiang, H. & Nie, D., 2011. Short-chain fatty acids induced autophagy serves as an adaptive strategy for retarding mitochondria-mediated apoptotic cell death. Cell Death and Differentiation, 2011(18), pp. 602-618. Watanabe, H. et al., 1993. cDNA Cloning of Transcriptiona Factor E4TF1 Subunits with Ets and Notch Motifs. Molecular and Cellualr Biology, 13(3), pp. 1385-1391. Wilkemeyer, M. et al., 1992. Propionate metabolism in cultured human cells after overexpression of recombinant methylmalonyl CoA mutase: Implications for somatic gene therapy. Somatic Cell and Molecular Genetics, 18(6), pp. 493-505. Wong, J. M. et al., 2006. Colonic Health: Fermentation and Short Chain Fatty Acids. Journal of Clinical Gastroenterology, 40(3), pp. 235-243. Read More