This web page was produced as an assignment for Genetics 564, an undergraduate course at UW-Madison
Determining the role of tubby in feeding behavior
Obesity
is a growing epidemic in the United States as over one-third of US adults are
currently obese. [1] In 2008, the US spent $147 million on
obesity related diseases including heart disease, stroke, type 2 diabetes, and
certain types of cancer. [1] One gene that has been shown to function in obesity is Tubby (Tub). The Tub gene was first
discovered in tubby mice that develop severe adult-onset obesity and insulin
resistance. [2, 3] TUB belongs to a family of tubby proteins
that include the four other members: TULP1, TULP2, TULP3, and TULP4. While the
tubby family proteins all contain a highly conserved tubby domain in their
C-terminus, TUB is the only family member that is associated with obesity. [4]
The
Tub gene is
highly expressed in the brain, particularly the hypothalamus, a region that is involved
in energy regulation. This localization in the brain has suggested that obesity
could be due to neuronal defects in the neuroendocrine control of feeding
behavior. [5,6]. Tubby mice consume more food and human
polymorphisms in the Tub gene also
correlate with increased carbohydrate intake in women. [7, 8] The
mechanism by which TUB regulates this feeding behavior on the molecular level remains
unknown. Hunger neuropeptides (NPY, POMC, AGRP, and Orexin) that function
to regulate feeding behavior are differentially expressed in tubby mice leading
to a current hypothesis that TUB
regulates the expression of hunger neuropeptides on a transcriptional level to
control feeding behavior. [9, 10]
The primary goal of this study is to determine how TUB regulates feeding behavior on the molecular level. I will identify the protein motifs in Tubby that are important for this behavior as well as DNA motifs in the hunger neuropeptides that Tubby regulates. A better understanding of how Tubby regulates feeding behavior will lead to potential drug targets in the treatment of obesity.
SPECIFIC AIM 1: Determine why mutations in TUB, but not TULPs, result in increased food intake.
Approach: Alignments using Clustal Omega, T-Coffee, and Muscle of TUB and TULP1-4 in both humans and mice will be used to identify conserved and non-conserved regions between Tubby family members. MEME will then be used to identify protein motifs within regions unique to TUB. All homologues will then be searched for the unique protein motifs identified in mice and humans.
Hypothesis: The protein motifs that are unique to TUB will be involved in TUB’s regulation of feeding behavior. This will be tested by mutating the identified motifs and observing feeding behavior.
SPECIFIC AIM 2: Identify DNA motifs in the differentially expressed hunger neuropeptides that are important for regulating food intake.
Approach: DREME will be used to identify DNA motifs that are common to all of the hunger neuropeptides that are differentially expressed in Tubby mice.
Hypothesis: TUB regulates the expression of hunger neuropeptides to control food intake by binding to the conserved DNA binding motifs. This will be tested using Restriction Endonuclease Protection Selection and Amplification (REPSA) with TUB and the identified hunger neuropeptide DNA binding motifs.
SPECIFIC AIM 3: Determine whether mutations in TUB affect food choice or food intake.
Approach: Tubby mice will be allowed to choose between high protein, carb, or fat diets and preference will be assessed by the amount of each food type consumed. Microarray will then be used to determine how food choice affects gene expression by assigning mice to a high protein, high carbohydrate, or high fat diet.
Hypothesis: Tubby mice will be more likely to choose high carbohydrate diets as compared to wild-type mice. Consumption of a high carbohydrate diet will affect the expression of TUB, its regulated hunger neuropeptides, and genes associated with metabolic processes and regulation according to Gene Ontology (GO) Biological Process classification.
The primary goal of this study is to determine how TUB regulates feeding behavior on the molecular level. I will identify the protein motifs in Tubby that are important for this behavior as well as DNA motifs in the hunger neuropeptides that Tubby regulates. A better understanding of how Tubby regulates feeding behavior will lead to potential drug targets in the treatment of obesity.
SPECIFIC AIM 1: Determine why mutations in TUB, but not TULPs, result in increased food intake.
Approach: Alignments using Clustal Omega, T-Coffee, and Muscle of TUB and TULP1-4 in both humans and mice will be used to identify conserved and non-conserved regions between Tubby family members. MEME will then be used to identify protein motifs within regions unique to TUB. All homologues will then be searched for the unique protein motifs identified in mice and humans.
Hypothesis: The protein motifs that are unique to TUB will be involved in TUB’s regulation of feeding behavior. This will be tested by mutating the identified motifs and observing feeding behavior.
SPECIFIC AIM 2: Identify DNA motifs in the differentially expressed hunger neuropeptides that are important for regulating food intake.
Approach: DREME will be used to identify DNA motifs that are common to all of the hunger neuropeptides that are differentially expressed in Tubby mice.
Hypothesis: TUB regulates the expression of hunger neuropeptides to control food intake by binding to the conserved DNA binding motifs. This will be tested using Restriction Endonuclease Protection Selection and Amplification (REPSA) with TUB and the identified hunger neuropeptide DNA binding motifs.
SPECIFIC AIM 3: Determine whether mutations in TUB affect food choice or food intake.
Approach: Tubby mice will be allowed to choose between high protein, carb, or fat diets and preference will be assessed by the amount of each food type consumed. Microarray will then be used to determine how food choice affects gene expression by assigning mice to a high protein, high carbohydrate, or high fat diet.
Hypothesis: Tubby mice will be more likely to choose high carbohydrate diets as compared to wild-type mice. Consumption of a high carbohydrate diet will affect the expression of TUB, its regulated hunger neuropeptides, and genes associated with metabolic processes and regulation according to Gene Ontology (GO) Biological Process classification.
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references
Cover Photo Credit
[1] Centers for Disease Control and Prevention: Adult Obesity Facts. <http://www.cdc.gov/obesity/data/adult.html>.
[2] North, M.A., Naggert, J.K., Yan, Y., Noben-Trauth, K., & Nishina, P.M. (1997). Molecular characterization of TUB, TULP1, and TULP2, members of the novel tubby gene family and their possible relation to ocular diseases. PNAS, 94(7), 3128.
[3] Coleman, D.L. & Eicher, E.M. (1990). Fat (fat) and Tubby (tub): Two Autosomal Recessive Mutations Causing Obesity Syndromes in the Mouse. Journal of Heredity, 81(1): 424-427.
[4] Mukhopadhyay and Jackson, 2011 The tubby family proteins Genome Biol. 2011 Jun 28;12(6):225. doi: 10.1186/gb-2011-12-6-225.
[5] Kleyn, P.W., Fan, W., Kovats, S. G., et al. (1996). Identification and Characterization of the Mouse Obesity Gene tubby: A Member of a Novel Gene Family. Cell, 85(2), doi:10.1016/S0092-8674(00)81104-6.
[6] Carroll, K., Gomez, C., & Shapiro, L. (2004). Tubby Proteins: The Plot Thickens. Molecular Cell Biology, 5(1), doi:10.1038/nrm1278.
[7] Backberg, M., Madjid, N., Ogren, S.O., and Meister, B. (2004). Down-regulated expression of agouti-related protein (AGRP) mRNA in the hypothalamic arcuate nucleus of hyperphagic and obese tub/tub mice. Molecular Brain Research, 125 (129). doi:10.1016/j.molbrainres.2004.03.012.
[8] Vliet-Ostaptchouk, J.V., Onland-Moret, N.C., Shiri-Sverdlov, R., et al. (2008). Polymorphisms of the TUB Gene Are Associated with Body Composition and Eating Behavior in Middle-Aged Women. PLoS ONE, 3(1), e1405. doi:10.1371/journal.pone.0001405.
[9] Guan, X.M., Yu, H., Lex, H.T., & Ploeg, V. (1998). Evidence of altered hypothalamic pro-opiomelancortin/neuropeptide Y mRNA expression in tubby mice. Molecular Brain Research, 59(273). doi: 10.1016/S0169328X98001508.
[10] Wang, Y., Seburn, K., Bechtel, L., Lee, B.Y., Szatkiewicz, J.P., Nishina, P.M., and Naggert, J.K. (2006). Defective carbohydrate metabolism in mice homozygous for the tubby mutation. Physiol Genomics, 27(131). doi:10.1152/physiolgenomics.00239.2005.
[1] Centers for Disease Control and Prevention: Adult Obesity Facts. <http://www.cdc.gov/obesity/data/adult.html>.
[2] North, M.A., Naggert, J.K., Yan, Y., Noben-Trauth, K., & Nishina, P.M. (1997). Molecular characterization of TUB, TULP1, and TULP2, members of the novel tubby gene family and their possible relation to ocular diseases. PNAS, 94(7), 3128.
[3] Coleman, D.L. & Eicher, E.M. (1990). Fat (fat) and Tubby (tub): Two Autosomal Recessive Mutations Causing Obesity Syndromes in the Mouse. Journal of Heredity, 81(1): 424-427.
[4] Mukhopadhyay and Jackson, 2011 The tubby family proteins Genome Biol. 2011 Jun 28;12(6):225. doi: 10.1186/gb-2011-12-6-225.
[5] Kleyn, P.W., Fan, W., Kovats, S. G., et al. (1996). Identification and Characterization of the Mouse Obesity Gene tubby: A Member of a Novel Gene Family. Cell, 85(2), doi:10.1016/S0092-8674(00)81104-6.
[6] Carroll, K., Gomez, C., & Shapiro, L. (2004). Tubby Proteins: The Plot Thickens. Molecular Cell Biology, 5(1), doi:10.1038/nrm1278.
[7] Backberg, M., Madjid, N., Ogren, S.O., and Meister, B. (2004). Down-regulated expression of agouti-related protein (AGRP) mRNA in the hypothalamic arcuate nucleus of hyperphagic and obese tub/tub mice. Molecular Brain Research, 125 (129). doi:10.1016/j.molbrainres.2004.03.012.
[8] Vliet-Ostaptchouk, J.V., Onland-Moret, N.C., Shiri-Sverdlov, R., et al. (2008). Polymorphisms of the TUB Gene Are Associated with Body Composition and Eating Behavior in Middle-Aged Women. PLoS ONE, 3(1), e1405. doi:10.1371/journal.pone.0001405.
[9] Guan, X.M., Yu, H., Lex, H.T., & Ploeg, V. (1998). Evidence of altered hypothalamic pro-opiomelancortin/neuropeptide Y mRNA expression in tubby mice. Molecular Brain Research, 59(273). doi: 10.1016/S0169328X98001508.
[10] Wang, Y., Seburn, K., Bechtel, L., Lee, B.Y., Szatkiewicz, J.P., Nishina, P.M., and Naggert, J.K. (2006). Defective carbohydrate metabolism in mice homozygous for the tubby mutation. Physiol Genomics, 27(131). doi:10.1152/physiolgenomics.00239.2005.
Site created by Rachael Baird.
Genetics 564 Assignment, Spring 2014
University of Wisconsin-Madison
Last Updated: 5-9-14
Genetics 564 Assignment, Spring 2014
University of Wisconsin-Madison
Last Updated: 5-9-14