Biomedical subjects

Audrey Robinson-White

Publications and source records attributed to Audrey Robinson-White.

7 recordsLinked to original sources

PubMed2006

Adrenal hyperplasia and adenomas are associated with inhibition of phosphodiesterase 11A in carriers of PDE11A sequence variants that are frequent in the population.

Several types of adrenocortical tumors that lead to Cushing syndrome may be caused by aberrant cyclic AMP (cAMP) signaling. We recently identified patients with micronodular adrenocortical hyperplasia who were carriers of inactivating mutations in the 2q-located phosphodiesterase 11A (PDE11A) gene. We now studied the frequency of two missense substitutions, R804H and R867G, in conserved regions of the enzyme in several sets of normal controls, including 745 individuals enrolled in a longitudinal cohort study, the New York Cancer Project. In the latter, we also screened for the presence of the previously identified PDE11A nonsense mutations. R804H and R867G were frequent among patients with adrenocortical tumors; although statistical significance was not reached, these variants affected significantly enzymatic function in vitro with variable increases in cAMP and/or cyclic guanosine 3',5'-monophosphate levels in HeLa and HEK293 cells. Adrenocortical tissues carrying the R804H mutation showed 2q allelic losses and higher cyclic nucleotide levels and cAMP-responsive element binding protein phosphorylation. We conclude that missense mutations of the PDE11A gene that affect enzymatic activity in vitro are present in the general population; protein-truncating PDE11A mutations may also contribute to a predisposition to other tumors, in addition to their association with adrenocortical hyperplasia. We speculate that PDE11A genetic defects may be associated with adrenal pathology in a wider than previously suspected clinical spectrum that includes asymptomatic individuals.

3',5'-Cyclic-GMP Phosphodiesterases↗

PubMed2006

A genome-wide scan identifies mutations in the gene encoding phosphodiesterase 11A4 (PDE11A) in individuals with adrenocortical hyperplasia.

Phosphodiesterases (PDEs) regulate cyclic nucleotide levels. Increased cyclic AMP (cAMP) signaling has been associated with PRKAR1A or GNAS mutations and leads to adrenocortical tumors and Cushing syndrome. We investigated the genetic source of Cushing syndrome in individuals with adrenocortical hyperplasia that was not caused by known defects. We performed genome-wide SNP genotyping, including the adrenocortical tumor DNA. The region with the highest probability to harbor a susceptibility gene by loss of heterozygosity (LOH) and other analyses was 2q31-2q35. We identified mutations disrupting the expression of the PDE11A isoform-4 gene (PDE11A) in three kindreds. Tumor tissues showed 2q31-2q35 LOH, decreased protein expression and high cyclic nucleotide levels and cAMP-responsive element binding protein (CREB) phosphorylation. PDE11A codes for a dual-specificity PDE that is expressed in adrenal cortex and is partially inhibited by tadalafil and other PDE inhibitors; its germline inactivation is associated with adrenocortical hyperplasia, suggesting another means by which dysregulation of cAMP signaling causes endocrine tumors.

3',5'-Cyclic-GMP Phosphodiesterases↗

PubMed2006

PRKAR1A Mutations and protein kinase A interactions with other signaling pathways in the adrenal cortex.

CONTEXT: Primary pigmented nodular adrenocortical disease, associated with Carney complex, is caused by mutations in PRKAR1A (mt-PRKAR1A), a gene that codes for the regulatory subunit type 1alpha (RIalpha) of cAMP-dependent protein kinase (PKA). PRKAR1A inactivation is associated with dysregulated PKA activity that is thought to result in tumorigenesis. mt-PRKAR1A-bearing lymphocytes from Carney complex patients exhibit enhanced cell proliferation associated with increased expression of the MAPK ERK1/2 pathway. OBJECTIVE: The objective of the study was to determine how PKA and its subunits and ERK1/2 and their molecular partners change in the presence of PRKAR1A mutations in adrenocortical tissue. DESIGN: PKA activity and subunit expression, ERK1/2, other immunoassays, and immunohistochemistry on adrenocortical samples from patients with germline normal or mt-PRKAR1A were analyzed. RESULTS: Increased cAMP-stimulated total kinase activity was associated with mt-PRKAR1A. PKA subunit expression analysis in mt-PRKAR1A tissues, by quantitative mRNA assay and immunoblotting, showed a 2.4-fold (P = 0.02) and 1.8-fold (P = 0.09) decrease in RIalpha's message and protein, respectively, and increases in other PKA subunits. Immunoassays showed 2-fold (P = 0.03) and 6-fold (P = 0.03) decreases in baseline ERK1/2, with corresponding increases in phosphorylated (p) ERK1/2 in mt-PRKAR1A samples. B-raf kinase, p-MEK1/2, and p-c-Myc, but not p-Akt/protein kinase B, were significantly increased. Immunohistochemistry studies supported these data. CONCLUSIONS: mt-PRKAR1A causes increased total cAMP-stimulated kinase activity, likely the result of up-regulation of other PKA subunits caused by down-regulation of RIalpha, as seen in human lymphocytes and mouse animal models. These changes, associated with enhanced MAPK activity, may be, in part, responsible for the proliferative signals that result in primary pigmented nodular adrenocortical disease.

Adrenal Cortex↗

PubMed2004

Down-regulation of regulatory subunit type 1A of protein kinase A leads to endocrine and other tumors.

Mutations of the human type Ialpha regulatory subunit (RIalpha) of cyclic AMP-dependent protein kinase (PKA; PRKAR1A) lead to altered kinase activity, primary pigmented nodular adrenocortical disease, and tumors of the thyroid and other tissues. To bypass the early embryonic lethality of Prkar1a(-/-) mice, we established transgenic mice carrying an antisense transgene for Prkar1a exon 2 (X2AS) under the control of a tetracycline-responsive promoter. Down-regulation of Prkar1a by up to 70% was achieved in transgenic mouse tissues and embryonic fibroblasts, with concomitant changes in kinase activity and increased cell proliferation, respectively. Mice developed thyroid follicular hyperplasia and adenomas, adrenocortical hyperplasia, and other features reminiscent of primary pigmented nodular adrenocortical disease, histiocytic and epithelial hyperplasias, lymphomas, and other mesenchymal tumors. These were associated with allelic losses of the mouse chromosome 11 Prkar1a locus, an increase in total type II PKA activity, and higher RIIbeta protein levels. This mouse provides a novel, useful tool for the investigation of cyclic AMP, RIalpha, and PKA functions and confirms the critical role of Prkar1a in tumorigenesis in endocrine and other tissues.

Adenoma↗

PubMed2004

A mouse model for Carney complex.

Mice with complete inactivation of the type Ialpha regulatory subunit (RIalpha) of cyclic (c) AMP-dependent protein kinase (PKA) (coded by the Prkar1a gene) die early in embryonic life. To bypass the early embryonic lethality of Prkar1a-/- mice, we established transgenic mice carrying an antisense transgene for Prkar1a exon 2 (X2AS) under the control of a tetracycline-responsive promoter. Mice developed thyroid follicular hyperplasia and adenomas, adrenocortical hyperplasia, and other features reminiscent of PPNAD, and histiocytic and epithelial hyperplasias, lymphomas, and other mesenchymal tumors. This mouse provides a useful tool for the investigation of cAMP, RIalpha, and PKA functions and confirms Prkar1a's critical role in tumorigenesis in endocrine and other tissues.

Animals↗

PubMed2003

Protein kinase-A activity in PRKAR1A-mutant cells, and regulation of mitogen-activated protein kinases ERK1/2.

Carney complex (CNC) is caused by PRKAR1A-inactivating mutations. PRKAR1A encodes the regulatory subunit type I-alpha (RIalpha) of the cAMP-dependent kinase (PKA) holoenzyme; how RIalpha insufficiency leads to tumorigenesis remains unclear. In many cells PKA inhibits the extracellular receptor kinase (ERK1/2) cascade of the mitogen-activated protein kinase (MAPK) pathway leading to inhibition of cell proliferation. We investigated whether the PKA-mediated inhibitory effect on ERK1/2 is affected in CNC cells that carry germline PRKAR1A mutations. PKA activity both at baseline and after stimulation with cAMP was augmented in cells carrying mutations. Quantitative message analysis showed that the main PKA subunits expressed were type I (RIalpha and RIbeta) but RIalpha was decreased in mutant cells. Immunoblot assays of ERK1/2 phosphorylation by the cell- and pathway-specific stimulant lysophosphatidic acid (LPA) showed activation of this pathway in a time- and concentration-dependent manner that was prevented by a specific inhibitor. There was a greater rate of growth in mutant cells; forskolin and isoproterenol inhibited LPA-induced ERK1/2 phosphorylation in normal but not in mutant cells. Forskolin inhibited LPA-induced cell proliferation and metabolism in normal cells, but stimulated these parameters in mutant cells. These data were also replicated in a pituitary tumor cell line carrying the most common PRKAR1A mutation (c.578del TG), and an in vitro construct of mutant PRKAR1A that was recently shown to lead to augmented PKA-mediated phosphorylation. We conclude that PKA activity in CNC cells is increased and that its stimulation by forskolin or isoproterenol increases LPA-induced ERK1/2 phosphorylation, cell metabolism and proliferation. Reversal of PKA-mediated inhibition of this MAPK pathway in CNC cells may contribute to tumorigenesis in this condition.

Animals↗

PubMed2002

Protein kinase A signaling: "cross-talk" with other pathways in endocrine cells.

Protein kinase A (PKA) signaling, in "classic" endocrine cell functioning, is known to mediate cAMP effects, generated through adenylate cyclase as a response to the activation of G protein-coupled receptors (GPCRs). This signaling system is highly versatile; its flexibility is supported by a number of adenylate cyclases, four PKA regulatory and three catalytic subunits, and several phosphodiesterases that close the negative feedback loop of cAMP generation, most molecules that are expressed in a tissue-specific manner. A central question, however, remains: how do the hundreds of GPCRs mediate their specific effects? Tissue specificity of the expression of the various components of the PKA system, albeit necessary, cannot be the only answer. It helps more to view PKA as a central hub that interacts with a variety of other signaling pathways in endocrine cells, not only mediating but also communicating cAMP effects to the mitogen-activated protein kinase (MAPK), protein kinase C and B (PKC and PKB/Akt, respectively). The net result of these complex interactions, evidence for which is reviewed in this chapter, is what we know as "cAMP effects." It is, perhaps, because of this complexity that investigations of PKA signaling in vivo and in vitro often give contradictory results and are difficult to interpret.

Animals↗