Caveat experimentor: reporting expression profiling data involving multiple, functionally redundant isoforms.
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Apoptosis↗
Biomedical subjects
R Brooks Robey
Publications and source records attributed to R Brooks Robey.
Regulation of mesangial cell hexokinase activity and expression by heparin-binding epidermal growth factor-like growth factor: epidermal growth factors and phorbol esters increase glucose metabolism via a common mechanism involving classic mitogen-activated protein kinase pathway activation and induction of hexokinase II expression.
Heparin-binding epidermal growth factor -like growth factor (HB-EGF) expression and hexokinase (HK) activity are increased in various pathologic renal conditions. Although the mitogenic properties of HB-EGF have been well characterized, its effects on glucose (Glc) metabolism have not. We therefore examined the possibility that HB-EGF might regulate HK activity and expression in glomerular mesangial cells, which constitute the principal renal cell type affected by a variety of pathologic conditions. Protein kinase C (PKC)-dependent classic mitogen-activated protein kinase (MAPK) pathway activation has been associated with increased HK activity in this cell type, so we also examined dependence upon these signaling intermediates. HB-EGF (> or =10 nm) increased total HK activity over 50% within 12-24 h, an effect mimicked by other EGF receptor agonists, but not by IGF-1 or elevated Glc. EGF receptor and classic MAPK pathway antagonists prevented this increase, as did general inhibitors of gene transcription and protein synthesis. Both HB-EGF and phorbol esters activated the classic MAPK pathway, albeit via PKC-independent and PKC-dependent mechanisms, respectively. Both stimuli were associated with increased HK activity, selectively increased HKII isoform expression, and increased Glc metabolism via both the glycolytic-tricarboxylic acid cycle route and the pentose phosphate pathway. HB-EGF thus constitutes a novel regulator of mesangial cell HK activity and Glc metabolism. HKII is the principal regulated isoform in these cells, as it is in insulin-sensitive peripheral tissues, such as muscle. However, the uniform requirement for classic MAPK pathway activation distinguishes HKII regulation in mesangial cells from that observed in muscle. These findings suggest a novel mechanism whereby growth factors may couple metabolism to glomerular injury.
Animals↗
LPA is a novel lipid regulator of mesangial cell hexokinase activity and HKII isoform expression.
The prototypical extracellular phospholipid mediator, lysophosphatidic acid (LPA), exhibits growth factor-like properties and represents an important survival factor in serum. This potent mesangial cell mitogen is increased in conditions associated with glomerular injury. It is also a known activator of the classic mitogen-activated protein kinase (MAPK) pathway, which plays an important role in the regulation of mesangial cell hexokinase (HK) activity. To better understand the mechanisms coupling metabolism to injury, we examined the ability of LPA to regulate HK activity and expression in cultured murine mesangial cells. LPA increased total HK activity in a concentration- and time-dependent manner, with maximal increases of >50% observed within 12 h of exposure to LPA concentrations > or =25 microM (apparent ED(50) 2 microM). These effects were associated with increased extracellular signal-regulated kinase (ERK) activity and were prevented by the pharmacological inhibition of either MAPK/ERK kinase or protein kinase C (PKC). Increased HK activity was also associated with increased glucose (Glc) utilization and lactate accumulation, as well as selectively increased HKII isoform abundance. The ability of exogenous LPA to increase HK activity was both Ca2+ independent and pertussis toxin insensitive and was mimicked by LPA-generating phospholipase A2. We conclude that LPA constitutes a novel lipid regulator of mesangial cell HK activity and Glc metabolism. This regulation requires sequential activation of both Ca2+-independent PKC and the classic MAPK pathway and culminates in increased HKII abundance. These previously unrecognized metabolic consequences of LPA stimulation have both physiological and pathophysiological implications. They also suggest a novel mechanism whereby metabolism may be coupled to cellular injury via extracellular lipid mediators.
Animals↗
A central role for Pyk2-Src interaction in coupling diverse stimuli to increased epithelial NBC activity.
Regulation of renal Na-HCO cotransporter (NBC1) activity by cholinergic agonists, ANG II, and acute acidosis (CO(2)) requires both Src family kinase (SFK) and classic MAPK pathway activation. The nonreceptor tyrosine kinase proline-rich tyrosine kinase 2 (Pyk2) couples discrete G protein-coupled receptor and growth factor receptor signaling to SFK activation. We examined the role of Pyk2-SFK interaction in coupling these stimuli to increased NBC1 activity in opossum kidney cells. Carbachol increased tyrosine autophosphorylation of endogenous Pyk2 and ectopically expressed wild-type Pyk2 and were abrogated by kinase-dead mutant (Pyk2-KD) overexpression. Pyk2 phosphorylation was calcium/calmodulin dependent, and Pyk2 associated with Src by means of SH2 domain interaction. Pyk2 phosphorylation and Pyk2-Src interaction by carbachol were mimicked by both ANG II and CO(2). To correlate Pyk2 autophosphorylation and Pyk2-Src interaction with NBC activity, cotransporter activity was measured in untransfected cells and in cells overexpressing Pyk2-KD in the presence or absence of carbachol, ANG II, or CO(2). In Pyk2-KD-overexpressing cells, the effect of carbachol, ANG II, and CO(2) was abolished. We conclude that Pyk2 plays a central role in coupling carbachol, ANG II, and CO(2) to increased NBC activity. This coupling is mediated by Pyk2 autophosphorylation and Pyk2-Src interaction.
Acidosis↗
Increased hexokinase activity, of either ectopic or endogenous origin, protects renal epithelial cells against acute oxidant-induced cell death.
Glucose (Glc) metabolism protects cells against oxidant injury. By virtue of their central position in both Glc uptake and utilization, hexokinases (HKs) are ideally suited to contribute to these effects. Compatible with this hypothesis, endogenous HK activity correlates inversely with injury susceptibility in individual renal cell types. We recently reported that ectopic HK expression mimics the anti-apoptotic effects of growth factors in cultured fibroblasts, but anti-apoptotic roles for HKs have not been examined in other cell types or in a cellular injury model. We therefore evaluated HK overexpression for the ability to mitigate acute oxidant-induced cell death in an established epithelial cell culture injury model. In parallel, we examined salutary heparin-binding epidermal growth factor (EGF)-like growth factor (HB-EGF) treatment for the ability to 1) increase endogenous HK activity and 2) mimic the protective effects of ectopic HK expression. Both HK overexpression and HB-EGF increased Glc-phosphorylating capacity and metabolism, and these changes were associated with markedly reduced susceptibility to acute oxidant-induced apoptosis. The uniform Glc dependence of these effects suggests an important adaptive role for Glc metabolism, and for HK activity in particular, in the promotion of epithelial cell survival. These findings also support the contention that HKs contribute to the protective effects of growth factors.
Adenoviridae↗