{"id":{"repo_id":"vt","oai_identifier":"oai:vtechworks.lib.vt.edu:10919/41882"},"canonical_url":"https://search.dev.ndltd.org/etd/vt/oai:vtechworks.lib.vt.edu:10919/41882","repository":{"repo_id":"vt","name":"Virginia Tech","base_url":"https://vtechworks.lib.vt.edu/oai/request"},"display":{"title":"Function of a C-rich region in the transcriptional regulation of the glycogen phosphorylase-2 gene in Dictyostelium discoideum","abstract":"The cellular slime mold Dictyostelium discoideum is a simple eukaryote that has been used as a model organism for the investigation of eukaryotic cell signaling, regulation of gene expression, and development. Transcription of the gp-2 gene is induced during development and is regulated by extracellular morphogens such as cyclic AMP (cAMP; Sucic et al., 1993) and differentiation induction factor (DIF; Yin et al., 1994a). These morphogens are known to be involved in regulating cellular differentiation and pattern formation in the multicellular development of Dictyostelium. This makes the gp-2 gene a good candidate for the investigation of the mechanisms of regulation of gene expression during cellular differentiation. The gp-2 gene has been cloned and previous analyses of the promoter with 5' deletions have revealed the presence of several regulatory regions which contain repeated sequence elements; TA-boxes, the TAG-boxes, and C-boxes (Sucic et al., 1993; Rutherford et al., submitted). The C-rich regulatory region contains two C-box repeats and is the most proximal of the regulatory regions. The aims of this investigation are the precise definition of regulatory elements within the C-rich regulatory region and identification of the role played by these sequences in the transcriptional regulation of the gp-2 gene. The effects of disrupting regulatory sequences within the C-rich region with upstream regulatory elements intact were investigated using internal deletions and site-directed mutations. Using the luciferase reporter gene system it was shown that site-directed mutation of the downstream C-box (CB-2) and adjacent bases results in a 50-fold decrease in the developmentally induced luciferase levels compared to a wild-type promoter construct (OS). An internal deletion of this mutant construct that deletes the upstream Cbox (CB-1) and intervening sequences has no further effect on levels of expression induced during development. These data suggest that CB-2 and adjacent sequences are the regulatory sequences within the C-rich region that are involved in the transcriptional regulation of gp-2. The mutant promoter constructs regulate gene expression during development with the same temporal profile as the wild-type construct indicating that these sequences are not involved with the timing of induction of expression of the gp-2 gene. The induction of the gp-2 promoter constructs by extracellular cAMP was analyzed. The wild-type construct was induced approximately 10-20-fold by extracellular cAMP. However, the site-directed mutated promoter construct was not induced by cAMP under the same conditions. This suggests that CB-2 and adjacent sequences are involved with the induction of gp-2 expression by extracellular cAMP. Previously, 5' deletion analyses have shown that upstream regulatory elements are involved in cAMP-responsive expression of the gp-2 gene (Sucic et al., 1993). The data presented here indicate that the regulation of expression of the gp-2 gene by extracellular cAMP also requires sequences in and around CB-2.","abstract_html":"The cellular slime mold Dictyostelium discoideum is a simple eukaryote that has been used as a model organism for the investigation of eukaryotic cell signaling, regulation of gene expression, and development. Transcription of the gp-2 gene is induced during development and is regulated by extracellular morphogens such as cyclic AMP (cAMP; Sucic et al., 1993) and differentiation induction factor (DIF; Yin et al., 1994a). These morphogens are known to be involved in regulating cellular differentiation and pattern formation in the multicellular development of Dictyostelium. This makes the gp-2 gene a good candidate for the investigation of the mechanisms of regulation of gene expression during cellular differentiation. The gp-2 gene has been cloned and previous analyses of the promoter with 5' deletions have revealed the presence of several regulatory regions which contain repeated sequence elements; TA-boxes, the TAG-boxes, and C-boxes (Sucic et al., 1993; Rutherford et al., submitted). The C-rich regulatory region contains two C-box repeats and is the most proximal of the regulatory regions. The aims of this investigation are the precise definition of regulatory elements within the C-rich regulatory region and identification of the role played by these sequences in the transcriptional regulation of the gp-2 gene. The effects of disrupting regulatory sequences within the C-rich region with upstream regulatory elements intact were investigated using internal deletions and site-directed mutations. Using the luciferase reporter gene system it was shown that site-directed mutation of the downstream C-box (CB-2) and adjacent bases results in a 50-fold decrease in the developmentally induced luciferase levels compared to a wild-type promoter construct (OS). An internal deletion of this mutant construct that deletes the upstream Cbox (CB-1) and intervening sequences has no further effect on levels of expression induced during development. These data suggest that CB-2 and adjacent sequences are the regulatory sequences within the C-rich region that are involved in the transcriptional regulation of gp-2. The mutant promoter constructs regulate gene expression during development with the same temporal profile as the wild-type construct indicating that these sequences are not involved with the timing of induction of expression of the gp-2 gene. The induction of the gp-2 promoter constructs by extracellular cAMP was analyzed. The wild-type construct was induced approximately 10-20-fold by extracellular cAMP. However, the site-directed mutated promoter construct was not induced by cAMP under the same conditions. This suggests that CB-2 and adjacent sequences are involved with the induction of gp-2 expression by extracellular cAMP. Previously, 5' deletion analyses have shown that upstream regulatory elements are involved in cAMP-responsive expression of the gp-2 gene (Sucic et al., 1993). The data presented here indicate that the regulation of expression of the gp-2 gene by extracellular cAMP also requires sequences in and around CB-2.","abstract_has_math":false,"creators":["Wu, Wen"],"institution":"Virginia Tech","degree_name":"Master of Science","degree_level":"masters","degree_discipline":"Biology","degree_department":"Biology","school":null,"contributors":[],"advisors":[],"committee_chairs":[],"committee_members":[],"year":1995,"date_issued":"1995","date_published":"1995","updated_at":"2026-07-22T22:20:11Z","subjects":[],"languages":["en"],"rights":["In Copyright"],"rights_urls":["http://rightsstatements.org/vocab/InC/1.0/"],"identifier_entries":[{"key":"dc:identifier.other","label":"Dc Identifier Other","values":["etd-03302010-020705"],"render_values":[{"text":"etd-03302010-020705","href":null,"code":true}]}]},"links":{"outbound_url":"http://hdl.handle.net/10919/41882","outbound_label":"Handle","outbound_source":"dc:identifier.uri"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:contributor.department","label":"Department","values":["Biology"]},{"key":"dc:creator","label":"Author","values":["Wu, Wen"]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:date.accessioned","label":"Dc Date Accessioned","values":["2014-03-14T21:32:45Z"]},{"key":"dc:date.available","label":"Dc Date Available","values":["2014-03-14T21:32:45Z","2010-03-30"]},{"key":"dc:date.issued","label":"Date","values":["1995"]},{"key":"dc:publisher","label":"Institution","values":["Virginia Tech"]},{"key":"dc:type","label":"Dc Type","values":["Thesis"]},{"key":"dc:type.dcmitype","label":"Dc Type Dcmitype","values":["Text"]},{"key":"thesis:degree_discipline","label":"Discipline","values":["Biology"]},{"key":"thesis:degree_level","label":"Degree Level","values":["masters"]},{"key":"thesis:degree_name","label":"Degree Name","values":["Master of Science"]},{"key":"thesis:institution_name","label":"Thesis Institution Name","values":["Virginia Polytechnic Institute and State University"]}]},{"id":"language_rights","label":"Language and Rights","entries":[{"key":"dc:language.iso","label":"Language (ISO)","values":["en"]},{"key":"dc:rights","label":"Dc Rights","values":["In Copyright"]},{"key":"dc:rights.uri","label":"Rights URI","values":["http://rightsstatements.org/vocab/InC/1.0/"]}]},{"id":"identifiers","label":"Identifiers","entries":[{"key":"dc:identifier.other","label":"Dc Identifier Other","values":["etd-03302010-020705"]},{"key":"dc:identifier.uri","label":"Identifier URI","values":["http://hdl.handle.net/10919/41882"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description.abstract","label":"Abstract","values":["The cellular slime mold Dictyostelium discoideum is a simple eukaryote that has been used as a model organism for the investigation of eukaryotic cell signaling, regulation of gene expression, and development. Transcription of the gp-2 gene is induced during development and is regulated by extracellular morphogens such as cyclic AMP (cAMP; Sucic et al., 1993) and differentiation induction factor (DIF; Yin et al., 1994a). These morphogens are known to be involved in regulating cellular differentiation and pattern formation in the multicellular development of Dictyostelium. This makes the gp-2 gene a good candidate for the investigation of the mechanisms of regulation of gene expression during cellular differentiation. The gp-2 gene has been cloned and previous analyses of the promoter with 5' deletions have revealed the presence of several regulatory regions which contain repeated sequence elements; TA-boxes, the TAG-boxes, and C-boxes (Sucic et al., 1993; Rutherford et al., submitted). The C-rich regulatory region contains two C-box repeats and is the most proximal of the regulatory regions. The aims of this investigation are the precise definition of regulatory elements within the C-rich regulatory region and identification of the role played by these sequences in the transcriptional regulation of the gp-2 gene. The effects of disrupting regulatory sequences within the C-rich region with upstream regulatory elements intact were investigated using internal deletions and site-directed mutations. Using the luciferase reporter gene system it was shown that site-directed mutation of the downstream C-box (CB-2) and adjacent bases results in a 50-fold decrease in the developmentally induced luciferase levels compared to a wild-type promoter construct (OS). An internal deletion of this mutant construct that deletes the upstream Cbox (CB-1) and intervening sequences has no further effect on levels of expression induced during development. These data suggest that CB-2 and adjacent sequences are the regulatory sequences within the C-rich region that are involved in the transcriptional regulation of gp-2. The mutant promoter constructs regulate gene expression during development with the same temporal profile as the wild-type construct indicating that these sequences are not involved with the timing of induction of expression of the gp-2 gene. The induction of the gp-2 promoter constructs by extracellular cAMP was analyzed. The wild-type construct was induced approximately 10-20-fold by extracellular cAMP. However, the site-directed mutated promoter construct was not induced by cAMP under the same conditions. This suggests that CB-2 and adjacent sequences are involved with the induction of gp-2 expression by extracellular cAMP. Previously, 5' deletion analyses have shown that upstream regulatory elements are involved in cAMP-responsive expression of the gp-2 gene (Sucic et al., 1993). The data presented here indicate that the regulation of expression of the gp-2 gene by extracellular cAMP also requires sequences in and around CB-2."]},{"key":"dc:description.degree","label":"Dc Description Degree","values":["Master of Science"]},{"key":"dc:format.medium","label":"Dc Format Medium","values":["BTD"]},{"key":"dc:format.mimetype","label":"Dc Format Mimetype","values":["application/pdf"]},{"key":"dc:title","label":"Title","values":["Function of a C-rich region in the transcriptional regulation of the glycogen phosphorylase-2 gene in Dictyostelium discoideum"]}]}],"canonical_facts":{"dc:contributor.department":["Biology"],"dc:creator":["Wu, Wen"],"dc:date.accessioned":["2014-03-14T21:32:45Z"],"dc:date.available":["2014-03-14T21:32:45Z","2010-03-30"],"dc:date.issued":["1995"],"dc:description.abstract":["The cellular slime mold Dictyostelium discoideum is a simple eukaryote that has been used as a model organism for the investigation of eukaryotic cell signaling, regulation of gene expression, and development. Transcription of the gp-2 gene is induced during development and is regulated by extracellular morphogens such as cyclic AMP (cAMP; Sucic et al., 1993) and differentiation induction factor (DIF; Yin et al., 1994a). These morphogens are known to be involved in regulating cellular differentiation and pattern formation in the multicellular development of Dictyostelium. This makes the gp-2 gene a good candidate for the investigation of the mechanisms of regulation of gene expression during cellular differentiation. The gp-2 gene has been cloned and previous analyses of the promoter with 5' deletions have revealed the presence of several regulatory regions which contain repeated sequence elements; TA-boxes, the TAG-boxes, and C-boxes (Sucic et al., 1993; Rutherford et al., submitted). The C-rich regulatory region contains two C-box repeats and is the most proximal of the regulatory regions. The aims of this investigation are the precise definition of regulatory elements within the C-rich regulatory region and identification of the role played by these sequences in the transcriptional regulation of the gp-2 gene. The effects of disrupting regulatory sequences within the C-rich region with upstream regulatory elements intact were investigated using internal deletions and site-directed mutations. Using the luciferase reporter gene system it was shown that site-directed mutation of the downstream C-box (CB-2) and adjacent bases results in a 50-fold decrease in the developmentally induced luciferase levels compared to a wild-type promoter construct (OS). An internal deletion of this mutant construct that deletes the upstream Cbox (CB-1) and intervening sequences has no further effect on levels of expression induced during development. These data suggest that CB-2 and adjacent sequences are the regulatory sequences within the C-rich region that are involved in the transcriptional regulation of gp-2. The mutant promoter constructs regulate gene expression during development with the same temporal profile as the wild-type construct indicating that these sequences are not involved with the timing of induction of expression of the gp-2 gene. The induction of the gp-2 promoter constructs by extracellular cAMP was analyzed. The wild-type construct was induced approximately 10-20-fold by extracellular cAMP. However, the site-directed mutated promoter construct was not induced by cAMP under the same conditions. This suggests that CB-2 and adjacent sequences are involved with the induction of gp-2 expression by extracellular cAMP. Previously, 5' deletion analyses have shown that upstream regulatory elements are involved in cAMP-responsive expression of the gp-2 gene (Sucic et al., 1993). The data presented here indicate that the regulation of expression of the gp-2 gene by extracellular cAMP also requires sequences in and around CB-2."],"dc:description.degree":["Master of Science"],"dc:format.medium":["BTD"],"dc:format.mimetype":["application/pdf"],"dc:identifier.other":["etd-03302010-020705"],"dc:identifier.uri":["http://hdl.handle.net/10919/41882"],"dc:language.iso":["en"],"dc:publisher":["Virginia Tech"],"dc:rights":["In Copyright"],"dc:rights.uri":["http://rightsstatements.org/vocab/InC/1.0/"],"dc:title":["Function of a C-rich region in the transcriptional regulation of the glycogen phosphorylase-2 gene in Dictyostelium discoideum"],"dc:type":["Thesis"],"dc:type.dcmitype":["Text"],"thesis:degree_discipline":["Biology"],"thesis:degree_level":["masters"],"thesis:degree_name":["Master of Science"],"thesis:institution_name":["Virginia Polytechnic Institute and State University"]},"updated_at":"2026-07-22T22:20:11Z"}