{"id":{"repo_id":"cambridge","oai_identifier":"oai:www.repository.cam.ac.uk:1810/373449"},"canonical_url":"https://search.dev.ndltd.org/etd/cambridge/oai:www.repository.cam.ac.uk:1810/373449","repository":{"repo_id":"cambridge","name":"Cambridge University","base_url":"https://api.repository.cam.ac.uk/server/oai/request"},"display":{"title":"From genomics to function: Canine genetics highlight novel mechanisms of obesity and related traits","abstract":"Obesity is an increasingly prevalent complex disease with burdens on both canine and human health. It is a multifactorial disease with many environmental and biological risk factors and is highly heritable in humans, pointing towards a role for genetics. Obesity heritability is also observed in dogs where breeds show different risk of obesity. Despite the negative health consequences that result from obesity in dogs, research into its causes is limited. However, thanks to the genetic architecture of dogs, shared environment between dogs and humans, and the shared risk factors and comorbidities, the canine model offers opportunity to study obesity. Investigations in this thesis aim to elucidate genetic influences in canine obesity and the functional mechanisms conferring these to better understand obesity in both dogs and humans. The hypothalamic melanocortin receptors (MCR) are central to the control of energy homeostasis across species. The first investigations in this thesis aimed to better understand the role of canine MCR genes in obesity. Missense variants MC3R p.M320I and MC4R p.V213F were detected at varying frequencies across the breed groups. The loss-of-function mutation, MC3R p.M320I, was common in Labrador retrievers. Carrying the mutation was associated with delayed onset of puberty and a decrease in adiposity and weight. The effect of MC3R p.M320I on weight was different in obese and non-obese dogs, supporting a role of MC3R in regulating the boundaries of weight. Carrying MC4R p.V213F had inconsistent results on adiposity and weight in different breeds. Evidence suggested this may be due to variable genetic background. Using *in vitro* signalling assays, the MC4R p.V213F variant was shown to have a small loss-of-function effect. Next, variants predicted to impact protein function within known hypothalamic genes were identified in beagles. These studies provide valuable insight into the hypothalamic control of energy homeostasis in canine obesity and its conserved roles between species. To better understand the genetic basis of obesity across different dog breeds, a genome wide association study (GWAS) for food motivation was performed in a large cohort of mixed ancestry dogs. Although no genome wide significant associations were identified, the nominally significant loci were narrow and six were prioritised. Relevant candidate genes were identified in these regions. This included the gene coding for ghrelin, a hormone known to regulate appetite. In addition, a region of high significance in this GWAS overlapped with one previously associated with breed average obesity risk. In this region was the R-spondin2 gene (RSPO2), well known in canine genetics for harbouring an insertion which causes the furnishings hair phenotype. Furnished dogs were identified as having an increased risk of diabetes, hypothesised to result from poor adipose expansion. To identify novel functional mechanisms of obesity, the results of an additional canine obesity GWAS were explored which had identified DENN domain containing 1B (*DENND1B*), a protein involved in trafficking. Cross-species evidence that genomic variation in human *DENND1B* alters expression in the hypothalamus suggested it may regulate MC4R signalling. *DENND1B* and *MC4R* were then shown to be expressed in the same hypothalamic cells using murine single cell RNA sequencing data. Subsequently, overexpression of DENND1B *in vitro*, showed a reduction in MC4R downstream signalling and an increase in receptor internalisation, consistent with an increase in obesity risk. This represents the discovery of a novel mechanism in the central regulation of energy homeostasis. DENND1B was also implicated in adipose function. Overexpression of DENND1B in murine pre-adipocytes resulted in reduced glucose uptake highlighting possible pleiotropic effects of DENND1B in obesity mechanisms. The results of this thesis showcase the benefits of studying complex disease in dogs and provide novel insights into the genetic basis of canine obesity. They also contribute to an improved understanding of the mechanisms important in energy homeostasis across species.","abstract_html":"Obesity is an increasingly prevalent complex disease with burdens on both canine and human health. It is a multifactorial disease with many environmental and biological risk factors and is highly heritable in humans, pointing towards a role for genetics. Obesity heritability is also observed in dogs where breeds show different risk of obesity. Despite the negative health consequences that result from obesity in dogs, research into its causes is limited. However, thanks to the genetic architecture of dogs, shared environment between dogs and humans, and the shared risk factors and comorbidities, the canine model offers opportunity to study obesity. Investigations in this thesis aim to elucidate genetic influences in canine obesity and the functional mechanisms conferring these to better understand obesity in both dogs and humans. The hypothalamic melanocortin receptors (MCR) are central to the control of energy homeostasis across species. The first investigations in this thesis aimed to better understand the role of canine MCR genes in obesity. Missense variants MC3R p.M320I and MC4R p.V213F were detected at varying frequencies across the breed groups. The loss-of-function mutation, MC3R p.M320I, was common in Labrador retrievers. Carrying the mutation was associated with delayed onset of puberty and a decrease in adiposity and weight. The effect of MC3R p.M320I on weight was different in obese and non-obese dogs, supporting a role of MC3R in regulating the boundaries of weight. Carrying MC4R p.V213F had inconsistent results on adiposity and weight in different breeds. Evidence suggested this may be due to variable genetic background. Using *in vitro* signalling assays, the MC4R p.V213F variant was shown to have a small loss-of-function effect. Next, variants predicted to impact protein function within known hypothalamic genes were identified in beagles. These studies provide valuable insight into the hypothalamic control of energy homeostasis in canine obesity and its conserved roles between species. To better understand the genetic basis of obesity across different dog breeds, a genome wide association study (GWAS) for food motivation was performed in a large cohort of mixed ancestry dogs. Although no genome wide significant associations were identified, the nominally significant loci were narrow and six were prioritised. Relevant candidate genes were identified in these regions. This included the gene coding for ghrelin, a hormone known to regulate appetite. In addition, a region of high significance in this GWAS overlapped with one previously associated with breed average obesity risk. In this region was the R-spondin2 gene (RSPO2), well known in canine genetics for harbouring an insertion which causes the furnishings hair phenotype. Furnished dogs were identified as having an increased risk of diabetes, hypothesised to result from poor adipose expansion. To identify novel functional mechanisms of obesity, the results of an additional canine obesity GWAS were explored which had identified DENN domain containing 1B (*DENND1B*), a protein involved in trafficking. Cross-species evidence that genomic variation in human *DENND1B* alters expression in the hypothalamus suggested it may regulate MC4R signalling. *DENND1B* and *MC4R* were then shown to be expressed in the same hypothalamic cells using murine single cell RNA sequencing data. Subsequently, overexpression of DENND1B *in vitro*, showed a reduction in MC4R downstream signalling and an increase in receptor internalisation, consistent with an increase in obesity risk. This represents the discovery of a novel mechanism in the central regulation of energy homeostasis. DENND1B was also implicated in adipose function. Overexpression of DENND1B in murine pre-adipocytes resulted in reduced glucose uptake highlighting possible pleiotropic effects of DENND1B in obesity mechanisms. The results of this thesis showcase the benefits of studying complex disease in dogs and provide novel insights into the genetic basis of canine obesity. They also contribute to an improved understanding of the mechanisms important in energy homeostasis across species.","abstract_has_math":false,"creators":["McClellan, Alyce"],"institution":"University of Cambridge","degree_name":"Doctor of Philosophy (PhD)","degree_level":"Doctoral","degree_discipline":null,"degree_department":null,"school":null,"contributors":[],"advisors":["Raffan, Eleanor"],"committee_chairs":[],"committee_members":[],"year":2024,"date_issued":"2024-02-02","date_published":"2024-02-02","updated_at":"2026-07-22T22:23:54Z","subjects":["Canine","DENND1B","Genetics","GWAS","MC3R","MC4R","Metabolism","Molecular genetics","Obesity"],"languages":["eng"],"rights":[],"rights_urls":["https://www.repository.cam.ac.uk/bitstreams/7c1ddf3f-8693-40b6-b13c-58bd6b40351d/download","https://www.rioxx.net/licenses/all-rights-reserved/"],"identifier_entries":[]},"links":{"outbound_url":"https://doi.org/10.17863/CAM.111851","outbound_label":"DOI","outbound_source":"dc:identifier.doi"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:contributor.advisor","label":"Advisor","values":["Raffan, Eleanor"]},{"key":"dc:contributor.sponsor","label":"Sponsor","values":["Darwin College David Ellar Scholarship"]},{"key":"dc:creator","label":"Author","values":["McClellan, Alyce"]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:date.issued","label":"Date","values":["2024-02-02"]},{"key":"dc:publisher.institution","label":"Dc Publisher Institution","values":["University of Cambridge"]},{"key":"dc:relation.isreferencedby.uri","label":"Dc Relation Isreferencedby URI","values":["https://www.repository.cam.ac.uk/handle/1810/373449"]},{"key":"dc:type","label":"Dc Type","values":["Thesis"]},{"key":"dc:type.qualificationlevel","label":"Dc Type Qualificationlevel","values":["Doctoral"]},{"key":"dc:type.qualificationname","label":"Dc Type Qualificationname","values":["Doctor of Philosophy (PhD)"]}]},{"id":"subjects_keywords","label":"Subjects and Keywords","entries":[{"key":"dc:subject","label":"Dc Subject","values":["Canine","DENND1B","Genetics","GWAS","MC3R","MC4R","Metabolism","Molecular genetics","Obesity"]}]},{"id":"language_rights","label":"Language and Rights","entries":[{"key":"dc:language","label":"Dc Language","values":["eng"]},{"key":"dc:rights","label":"Dc Rights","values":["https://www.repository.cam.ac.uk/bitstreams/7c1ddf3f-8693-40b6-b13c-58bd6b40351d/download","https://www.rioxx.net/licenses/all-rights-reserved/"]}]},{"id":"identifiers","label":"Identifiers","entries":[{"key":"dc:identifier.doi","label":"DOI","values":["https://doi.org/10.17863/CAM.111851"]},{"key":"dc:identifier.uri","label":"Identifier URI","values":["https://www.repository.cam.ac.uk/bitstreams/f0b40eef-39b0-4307-933e-c918cf6093d0/download"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description.abstract","label":"Abstract","values":["Obesity is an increasingly prevalent complex disease with burdens on both canine and human health. 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Missense variants MC3R p.M320I and MC4R p.V213F were detected at varying frequencies across the breed groups. The loss-of-function mutation, MC3R p.M320I, was common in Labrador retrievers. Carrying the mutation was associated with delayed onset of puberty and a decrease in adiposity and weight. The effect of MC3R p.M320I on weight was different in obese and non-obese dogs, supporting a role of MC3R in regulating the boundaries of weight. Carrying MC4R p.V213F had inconsistent results on adiposity and weight in different breeds. Evidence suggested this may be due to variable genetic background. Using *in vitro* signalling assays, the MC4R p.V213F variant was shown to have a small loss-of-function effect. Next, variants predicted to impact protein function within known hypothalamic genes were identified in beagles. These studies provide valuable insight into the hypothalamic control of energy homeostasis in canine obesity and its conserved roles between species. To better understand the genetic basis of obesity across different dog breeds, a genome wide association study (GWAS) for food motivation was performed in a large cohort of mixed ancestry dogs. Although no genome wide significant associations were identified, the nominally significant loci were narrow and six were prioritised. Relevant candidate genes were identified in these regions. This included the gene coding for ghrelin, a hormone known to regulate appetite. In addition, a region of high significance in this GWAS overlapped with one previously associated with breed average obesity risk. In this region was the R-spondin2 gene (RSPO2), well known in canine genetics for harbouring an insertion which causes the furnishings hair phenotype. Furnished dogs were identified as having an increased risk of diabetes, hypothesised to result from poor adipose expansion. To identify novel functional mechanisms of obesity, the results of an additional canine obesity GWAS were explored which had identified DENN domain containing 1B (*DENND1B*), a protein involved in trafficking. Cross-species evidence that genomic variation in human *DENND1B* alters expression in the hypothalamus suggested it may regulate MC4R signalling. *DENND1B* and *MC4R* were then shown to be expressed in the same hypothalamic cells using murine single cell RNA sequencing data. Subsequently, overexpression of DENND1B *in vitro*, showed a reduction in MC4R downstream signalling and an increase in receptor internalisation, consistent with an increase in obesity risk. This represents the discovery of a novel mechanism in the central regulation of energy homeostasis. DENND1B was also implicated in adipose function. Overexpression of DENND1B in murine pre-adipocytes resulted in reduced glucose uptake highlighting possible pleiotropic effects of DENND1B in obesity mechanisms. 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Missense variants MC3R p.M320I and MC4R p.V213F were detected at varying frequencies across the breed groups. The loss-of-function mutation, MC3R p.M320I, was common in Labrador retrievers. Carrying the mutation was associated with delayed onset of puberty and a decrease in adiposity and weight. The effect of MC3R p.M320I on weight was different in obese and non-obese dogs, supporting a role of MC3R in regulating the boundaries of weight. Carrying MC4R p.V213F had inconsistent results on adiposity and weight in different breeds. Evidence suggested this may be due to variable genetic background. Using *in vitro* signalling assays, the MC4R p.V213F variant was shown to have a small loss-of-function effect. Next, variants predicted to impact protein function within known hypothalamic genes were identified in beagles. These studies provide valuable insight into the hypothalamic control of energy homeostasis in canine obesity and its conserved roles between species. To better understand the genetic basis of obesity across different dog breeds, a genome wide association study (GWAS) for food motivation was performed in a large cohort of mixed ancestry dogs. Although no genome wide significant associations were identified, the nominally significant loci were narrow and six were prioritised. Relevant candidate genes were identified in these regions. This included the gene coding for ghrelin, a hormone known to regulate appetite. In addition, a region of high significance in this GWAS overlapped with one previously associated with breed average obesity risk. In this region was the R-spondin2 gene (RSPO2), well known in canine genetics for harbouring an insertion which causes the furnishings hair phenotype. Furnished dogs were identified as having an increased risk of diabetes, hypothesised to result from poor adipose expansion. To identify novel functional mechanisms of obesity, the results of an additional canine obesity GWAS were explored which had identified DENN domain containing 1B (*DENND1B*), a protein involved in trafficking. Cross-species evidence that genomic variation in human *DENND1B* alters expression in the hypothalamus suggested it may regulate MC4R signalling. *DENND1B* and *MC4R* were then shown to be expressed in the same hypothalamic cells using murine single cell RNA sequencing data. Subsequently, overexpression of DENND1B *in vitro*, showed a reduction in MC4R downstream signalling and an increase in receptor internalisation, consistent with an increase in obesity risk. This represents the discovery of a novel mechanism in the central regulation of energy homeostasis. DENND1B was also implicated in adipose function. Overexpression of DENND1B in murine pre-adipocytes resulted in reduced glucose uptake highlighting possible pleiotropic effects of DENND1B in obesity mechanisms. 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