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Cloning and quantitative determination of the human Ca2+/calmodulin-dependent protein kinase II (CaMK II) isoforms in
H Rochlitz1, A Voigt, B Lankat-Buttgereit
1Department of Internal Medicine, University Hospital Bergmannsheil, University of Bochum, Germany.
Aims/Hypothesis:
The Ca2+/calmodulin-dependent protein kinase II (CaMK II) is highly expressed in pancreatic islets and associated with insulin secretion vesicles. The suppression of CaMK II disturbs insulin secretion and insulin gene expression. There are four isoforms of CaMK II, alpha to delta, that are expressed from different genes in mammals. Our aim was to identify the isoforms of CaMK II expressed in human beta cells by molecular cloning from a human insulinoma cDNA library and to assess its distribution in humans.
Methods:
The previously unknown complete coding sequences of human CaMK IIbeta and the kinase domain of CaMK IIdelta were cloned from a human insulinoma cDNA library. Quantitative determination of CaMK II isoform mRNA was carried out in several tissues and beta cells purified by fluorescence activated cell sorting and compared to the housekeeping enzyme pyruvate dehydrogenase.
Results:
We found CaMK IIbeta occurred in three splice variants and was highly expressed in endocrine tissues such as adrenals, pituitary and beta cells. Liver showed moderate expression but adipose tissue or lymphocytes had very low levels of CaMK IIbeta-mRNA. In human beta cells CaMK IIbeta and delta were expressed equally with pyruvate dehydrogenase whereas tenfold lower expression of CaMK IIgamma and no expression of CaMK IIalpha were found.
Conclusion/Interpretation:
Although CaMK IIdelta is ubiquitously expressed, CaMK IIbeta shows preferential expression in neuroendocrine tissues. In comparison with the expression of a key regulatory enzyme in glucose oxidation, pyruvate dehydrogenase, two of the four CaM kinases investigated are expressed at equally high levels, which supports an important role in beta-cell physiology. These results provide the basis for exploring the pathophysiological relevance of CaMK IIbeta in human diabetes.
Insights
Calcium/calmodulin-dependent protein kinase II (CaMK II) beta and delta isoforms are highly expressed in human beta cells, suggesting a key role in insulin secretion and potential relevance to diabetes.
Area of Science:
- Molecular biology
- Endocrinology
- Cell biology
Background:
- Calcium/calmodulin-dependent protein kinase II (CaMK II) is crucial for insulin secretion and gene expression in pancreatic islets.
- Four mammalian CaMK II isoforms (alpha, beta, gamma, delta) exist, originating from distinct genes.
Purpose of the Study:
- To identify CaMK II isoforms in human beta cells using molecular cloning.
- To determine the distribution of CaMK II isoforms in various human tissues.
Main Methods:
- Molecular cloning of human CaMK IIbeta and CaMK IIdelta from an insulinoma cDNA library.
- Quantitative mRNA analysis of CaMK II isoforms in diverse tissues and purified beta cells.
- Comparison of CaMK II expression levels with the housekeeping enzyme pyruvate dehydrogenase.
Main Results:
- CaMK IIbeta was found in three splice variants and showed high expression in endocrine tissues, including human beta cells.
- Human beta cells equally expressed CaMK IIbeta and CaMK IIdelta, with lower CaMK IIgamma and no CaMK IIalpha expression.
- CaMK IIbeta mRNA levels were high in adrenals and pituitary but low in liver, adipose tissue, and lymphocytes.
Conclusions:
- CaMK IIbeta exhibits preferential expression in neuroendocrine tissues, contrasting with the ubiquitous distribution of CaMK IIdelta.
- High expression levels of CaMK IIbeta and delta in beta cells, relative to pyruvate dehydrogenase, underscore their physiological importance.
- These findings establish a foundation for investigating the role of CaMK IIbeta in human diabetes pathophysiology.