Mice with a disruption in the hoxb-2 locus were generated by gene targeting. 75% of the hoxb-2 mu... more Mice with a disruption in the hoxb-2 locus were generated by gene targeting. 75% of the hoxb-2 mutant homozygotes died within 24 hours of birth. While a majority of these mice had severe sternal defects that compromised their ability to breathe, some had relatively normal sternum morphology, suggesting that one or more additional factor(s) contributed to neonatal lethality. At 3–3.5 weeks of age, half of the remaining hoxb-2 homozygotes became weak and subsequently died. All of the mutants that survived to 3 weeks of age showed marked facial paralysis similar to, but more severe than, that reported for hoxb-1 mutant homozygotes (Goddard, J. M., Rossel, M., Manley, N. R. and Capecchi, M. R. (1996) Development 122, 3217–3228). As for the hoxb-1 mutations, the facial paralysis observed in mice homozygous for the hoxb-2 mutation results from a failure to form the somatic motor component of the VIIth (facial) nerve which controls the muscles of facial expression. Features of this phenotype closely resemble the clinical signs associated with Bell's Palsy and Moebius Syndrome in humans. The sternal defects seen in hoxb-2 mutant mice are similar to those previously reported for hoxb-4 mutant mice (Ramirez-Solis, R., Zheng, H., Whiting, J., Krumlauf, R. and Bradley. A. (1993) Cell 73, 279–294). The above results suggest that the hoxb-2 mutant phenotype may result in part from effects of the hoxb-2 mutation on the expression of both hoxb-1 and hoxb-4. Consistent with this proposal, we found that the hoxb-2 mutation disrupts the expression of hoxb-1 in cis. In addition, the hoxb-2 mutation changes the expression of hoxb-4 and the hoxb-4 mutation, in turn, alters the pattern of hoxb-2 expression. Hoxb-2 and hoxb-4 appear to function together to mediate proper closure of the ventral thoracic body wall. Failure in this closure results in severe defects of the sternum.
AMP-activated protein kinase (AMPK) has been identified as a regulator of gene transcription, inc... more AMP-activated protein kinase (AMPK) has been identified as a regulator of gene transcription, increasing mitochondrial proteins of oxidative metabolism as well as hexokinase expression in skeletal muscle. In mice, muscle-specific knockout of LKB1, a component of the upstream kinase of AMPK, prevents contraction- and 5-aminoimidazole-4-carboxamide-1-β-D-ribofuranoside (AICAR)-induced activation of AMPK in skeletal muscle, and the increase in hexokinase II protein that is normally observed with chronic AICAR activation of AMPK. Since previous reports show a cAMP response element in the promoter region of the hexokinase II gene, we hypothesized that the cAMP-response element (CRE) binding protein (CREB) family of transcription factors could be targets of AMPK. Using radioisotopic kinase assays, we found that recombinant and rat liver and muscle AMPK phosphorylated CREB1 at the same site as cAMP-dependent protein kinase (PKA). AMPK was also found to phosphorylate activating transcriptio...
This Thesis is brought to you for free and open access by BYU ScholarsArchive. It has been accept... more This Thesis is brought to you for free and open access by BYU ScholarsArchive. It has been accepted for inclusion in All Theses and Dissertations by an authorized administrator of BYU ScholarsArchive. For more information, please contact scholarsarchive@byu.edu.
This Dissertation is brought to you for free and open access by BYU ScholarsArchive. It has been ... more This Dissertation is brought to you for free and open access by BYU ScholarsArchive. It has been accepted for inclusion in All Theses and Dissertations by an authorized administrator of BYU ScholarsArchive. For more information, please contact scholarsarchive@byu.edu.
Tissue Specific Porcupine Deletion Reveals a Novel Role for Ectodermal Wnts in Musculotendon Deve... more Tissue Specific Porcupine Deletion Reveals a Novel Role for Ectodermal Wnts in Musculotendon Development Aaron P. Smith Department of Physiology and Developmental Biology, BYU Masters of Science The Wnt family of secreted proteins consists of 19 family members (in the mouse) and is known to signal through multiple pathways that regulate crucial processes in the development of almost all tissues. Dissecting the roles of individual Wnts has been hampered due to functional redundancy that exists between family members. We made use of a conditional allele of the Oacyltransferase, Porcupine (Porcn), that is required for the secretion of all Wnt ligands, and the Msx2Cre deleter to eliminate the secretion of all Wnt ligands from the ventral limb ectoderm, ventral abdominal ectoderm, and urogenital ectoderm. Phenotypically the limbs of these mice have several similarities with En1 mutant mice which have a double-dorsal phenotype. However, we show that appropriate dorsoventral limb pattern i...
CONSTRUCTION OF A COL11A1 TRANSGENE VECTOR Cameron McKell Beck Department of Physiology and Devel... more CONSTRUCTION OF A COL11A1 TRANSGENE VECTOR Cameron McKell Beck Department of Physiology and Developmental Biology Master of Science Background: Cartilage disorders affect millions of people in the United States alone, with effects ranging from poor skeletal development and joint pain to shortened lifespan and perinatal lethality. Many of these disorders have their root in defects of collagen, type XI collagen being among the most important. A mouse model of such a type XI collagen defect is the chondrodysplasia (cho) mutant. Mice homozygous for this null mutation in the Col11a1 gene do not express the α1 chain of type XI collagen. This results in a functional knockout of type XI collagen, leading to insufficient skeletal development and perinatal lethality. Objective: 1) To construct a transgenic expression vector designed to express a human COL11A1 cDNA in a cartilage-specific manner. This transgene will be used in future studies to correct the type XI collagen defect in homozygous...
Role of Wnt5a and Possible Pathway of Action through Ror2 in Proximodistal Outgrowth of the Limb ... more Role of Wnt5a and Possible Pathway of Action through Ror2 in Proximodistal Outgrowth of the Limb Tiffanie M. Dahl Department of Physiology and Developmental Biology, BYU Master of Science Despite over 60 years of study, the molecular pathways and mechanisms governing limb outgrowth and patterning remain poorly understood. Fgfs expressed in the AER are known to be necessary and sufficient for proximodistal limb outgrowth and have been proposed to have a chemoattractive role. Wnt5a is a secreted factor which is expressed in a gradient in the distal limb with the highest concentration next to the AER. The presence of the AER is necessary to establish this gradient. Expression of Wnt5a in a concentration dependant manner can be induced in the limb through the implantation of a bead soaked in recombinant Fgf4 protein. This indicates that Fgfs from the AER may establish the gradient of Wnt5a in the limb mesenchyme. Wnt5a mutants exhibit severe shortening of the face, limbs, and body axis,...
Mice with a disruption in the hoxb-2 locus were generated by gene targeting. 75% of the hoxb-2 mu... more Mice with a disruption in the hoxb-2 locus were generated by gene targeting. 75% of the hoxb-2 mutant homozygotes died within 24 hours of birth. While a majority of these mice had severe sternal defects that compromised their ability to breathe, some had relatively normal sternum morphology, suggesting that one or more additional factor(s) contributed to neonatal lethality. At 3–3.5 weeks of age, half of the remaining hoxb-2 homozygotes became weak and subsequently died. All of the mutants that survived to 3 weeks of age showed marked facial paralysis similar to, but more severe than, that reported for hoxb-1 mutant homozygotes (Goddard, J. M., Rossel, M., Manley, N. R. and Capecchi, M. R. (1996) Development 122, 3217–3228). As for the hoxb-1 mutations, the facial paralysis observed in mice homozygous for the hoxb-2 mutation results from a failure to form the somatic motor component of the VIIth (facial) nerve which controls the muscles of facial expression. Features of this phenotype closely resemble the clinical signs associated with Bell's Palsy and Moebius Syndrome in humans. The sternal defects seen in hoxb-2 mutant mice are similar to those previously reported for hoxb-4 mutant mice (Ramirez-Solis, R., Zheng, H., Whiting, J., Krumlauf, R. and Bradley. A. (1993) Cell 73, 279–294). The above results suggest that the hoxb-2 mutant phenotype may result in part from effects of the hoxb-2 mutation on the expression of both hoxb-1 and hoxb-4. Consistent with this proposal, we found that the hoxb-2 mutation disrupts the expression of hoxb-1 in cis. In addition, the hoxb-2 mutation changes the expression of hoxb-4 and the hoxb-4 mutation, in turn, alters the pattern of hoxb-2 expression. Hoxb-2 and hoxb-4 appear to function together to mediate proper closure of the ventral thoracic body wall. Failure in this closure results in severe defects of the sternum.
AMP-activated protein kinase (AMPK) has been identified as a regulator of gene transcription, inc... more AMP-activated protein kinase (AMPK) has been identified as a regulator of gene transcription, increasing mitochondrial proteins of oxidative metabolism as well as hexokinase expression in skeletal muscle. In mice, muscle-specific knockout of LKB1, a component of the upstream kinase of AMPK, prevents contraction- and 5-aminoimidazole-4-carboxamide-1-β-D-ribofuranoside (AICAR)-induced activation of AMPK in skeletal muscle, and the increase in hexokinase II protein that is normally observed with chronic AICAR activation of AMPK. Since previous reports show a cAMP response element in the promoter region of the hexokinase II gene, we hypothesized that the cAMP-response element (CRE) binding protein (CREB) family of transcription factors could be targets of AMPK. Using radioisotopic kinase assays, we found that recombinant and rat liver and muscle AMPK phosphorylated CREB1 at the same site as cAMP-dependent protein kinase (PKA). AMPK was also found to phosphorylate activating transcriptio...
This Thesis is brought to you for free and open access by BYU ScholarsArchive. It has been accept... more This Thesis is brought to you for free and open access by BYU ScholarsArchive. It has been accepted for inclusion in All Theses and Dissertations by an authorized administrator of BYU ScholarsArchive. For more information, please contact scholarsarchive@byu.edu.
This Dissertation is brought to you for free and open access by BYU ScholarsArchive. It has been ... more This Dissertation is brought to you for free and open access by BYU ScholarsArchive. It has been accepted for inclusion in All Theses and Dissertations by an authorized administrator of BYU ScholarsArchive. For more information, please contact scholarsarchive@byu.edu.
Tissue Specific Porcupine Deletion Reveals a Novel Role for Ectodermal Wnts in Musculotendon Deve... more Tissue Specific Porcupine Deletion Reveals a Novel Role for Ectodermal Wnts in Musculotendon Development Aaron P. Smith Department of Physiology and Developmental Biology, BYU Masters of Science The Wnt family of secreted proteins consists of 19 family members (in the mouse) and is known to signal through multiple pathways that regulate crucial processes in the development of almost all tissues. Dissecting the roles of individual Wnts has been hampered due to functional redundancy that exists between family members. We made use of a conditional allele of the Oacyltransferase, Porcupine (Porcn), that is required for the secretion of all Wnt ligands, and the Msx2Cre deleter to eliminate the secretion of all Wnt ligands from the ventral limb ectoderm, ventral abdominal ectoderm, and urogenital ectoderm. Phenotypically the limbs of these mice have several similarities with En1 mutant mice which have a double-dorsal phenotype. However, we show that appropriate dorsoventral limb pattern i...
CONSTRUCTION OF A COL11A1 TRANSGENE VECTOR Cameron McKell Beck Department of Physiology and Devel... more CONSTRUCTION OF A COL11A1 TRANSGENE VECTOR Cameron McKell Beck Department of Physiology and Developmental Biology Master of Science Background: Cartilage disorders affect millions of people in the United States alone, with effects ranging from poor skeletal development and joint pain to shortened lifespan and perinatal lethality. Many of these disorders have their root in defects of collagen, type XI collagen being among the most important. A mouse model of such a type XI collagen defect is the chondrodysplasia (cho) mutant. Mice homozygous for this null mutation in the Col11a1 gene do not express the α1 chain of type XI collagen. This results in a functional knockout of type XI collagen, leading to insufficient skeletal development and perinatal lethality. Objective: 1) To construct a transgenic expression vector designed to express a human COL11A1 cDNA in a cartilage-specific manner. This transgene will be used in future studies to correct the type XI collagen defect in homozygous...
Role of Wnt5a and Possible Pathway of Action through Ror2 in Proximodistal Outgrowth of the Limb ... more Role of Wnt5a and Possible Pathway of Action through Ror2 in Proximodistal Outgrowth of the Limb Tiffanie M. Dahl Department of Physiology and Developmental Biology, BYU Master of Science Despite over 60 years of study, the molecular pathways and mechanisms governing limb outgrowth and patterning remain poorly understood. Fgfs expressed in the AER are known to be necessary and sufficient for proximodistal limb outgrowth and have been proposed to have a chemoattractive role. Wnt5a is a secreted factor which is expressed in a gradient in the distal limb with the highest concentration next to the AER. The presence of the AER is necessary to establish this gradient. Expression of Wnt5a in a concentration dependant manner can be induced in the limb through the implantation of a bead soaked in recombinant Fgf4 protein. This indicates that Fgfs from the AER may establish the gradient of Wnt5a in the limb mesenchyme. Wnt5a mutants exhibit severe shortening of the face, limbs, and body axis,...
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