RC3 mRNA Research Articles

Cell-specific effects of thyroid hormone on RC3/neurogranin expression in rat brain

To identify thyroid hormone-sensitive neuronal populations in the forebrain, we studied the effects of thyroid hormone deficiency and replacement on the expression of RC3 messenger RNA (mRNA) in the rat brain by in situ hybridization. RC3/neurogranin is a brain-specific, calmodulin-binding, protein kinase C substrate that has been implicated in postsynaptic events involving calcium as a second messenger. We have previously shown that RC3 mRNA and protein concentrations are thyroid hormone dependent in developing and adult rats. In normal developing rats, RC3 expression occurs in two phases. Before postnatal day 10 (P10), RC3 mRNA was detected mainly in layers II/III and V of cerebral cortex and the CA fields of the hippocampus. From P10 to P15, it decreased in layer V and increased in layer VI, the retrosplenial cortex, the caudate-putamen nucleus, and the dentate gyrus. Expression in the caudate followed a lateral to medial gradient. Thyroid hormone deficiency interfered with the late phase of RC3 expression, such that developing hypothyroid rats showed lower RC3 expression in layer VI, the retrosplenial cortex, the dentate gyrus, and the caudate, and increased expression in layer V. These changes were reverted by T4 treatment. Adult- onset hyperthyroidism also reversibly decreased hybridization in the striatum. In contrast to other molecular targets of thyroid hormone in the brain, such as myelin genes, expression of RC3 was also affected by long term hypothyroidism in the absence of hormone replacement, indicating that thyroid hormone is a required factor for the cell-specific control of RC3 expression. In addition to identifying thyroid hormone-sensitive neurons, our results suggest that one action of thyroid hormone during brain development is the timely coordination of gene expression among phenotypically different, region-specific neuronal populations.

RC3/neurogranin structure and expression in the caprine brain in relation to congenital hypothyroidism

In view of the profound effects of thyroid hormone deficiency on the central nervous system (CNS), neuronal genes regulated by thyroid hormone could potentially be involved in the development of the CNS. Expression of the neuronal gene RC3/neurogranin was shown to be induced by thyroid hormone in the rat. No data are available on RC3 expression in mammals with prenatal brain development, like humans. To study RC3 mRNA expression in a genetic in vivo model of congenital hypothyroidism, which also resembles the human situation in the timing of brain development relative to birth, we used an inbred strain of congenitally hypothyroid goats. We isolated a cDNA for the caprine RC3 homolog. The deduced amino acid sequence had 99% similarity with the rat and bovine protein sequence. An analysis of the developmental expression of RC3 mRNA levels showed a 3-fold increase between E90 and P0. In situ hybridization analysis showed that in euthyroid goats, the RC3 expression pattern was region-specific and resembled that in rats. However, in contrast to rats, hypothyroid goats showed only a reduced RC3 mRNA expression in the striatum. Hypothyroidism had no effect on RC3 mRNA expression in all other brain regions. T4-treatment of the hypothyroid fetus increased RC3 mRNA expression in the striatum to euthyroid control levels. These data suggest that thyroid hormone is a regulator of RC3 gene expression in the caprine brain, and that the striatum is highly sensitive to thyroid hormone deficiency.

Adult rat brain is sensitive to thyroid hormone. Regulation of RC3/neurogranin mRNA.

The mammalian brain is considered to be poorly responsive to thyroid hormone after the so called "critical periods" of brain development, which occur in the rat before postnatal days 15-20. In a previous work (Muñoz, A., A. Rodriguez-Peña, A. Perez-Castillo, B. Ferreiro, J.G. Sutcliffe, and J. Bernal. 1991. Mol. Endocrinol. 5:273-280) we have identified one neuronal gene, RC3, whose expression is influenced by early neonatal hypothyroidism and thyroid hormone treatment. In the present work we show that adult-onset hypothyroidism leads to a reversible decrease of RC3 mRNA. Rats thyroidectomized on postnatal day 40 and killed three months later showed a decreased RC3 mRNA concentration in the cerebral cortex and striatum. The same effect was observed in animals made hypothyroid on postnatal day 32 and killed on postnatal day 52. RC3 expression was normal when hypothyroid animals were treated with T4 five days before being killed. In contrast, the mRNA encoding myelin proteolipid protein showed no changes in either experimental situation. RC3 mRNA levels were not affected by food restriction demonstrating that the effect of hypothyroidism was not related to the lack of weight gain. The control of RC3 mRNA is so far the only molecular event known to be regulated by thyroid hormone once the critical periods of brain development are over and could represent a molecular correlate for the age-independent, reversible alterations induced by hypothyroidism in the adult brain.

Subtractive cDNA cloning of RC3, a rodent cortex‐enriched mRNA encoding a novel 78 residue protein

A rodent cortex-enriched mRNA, RC3, was identified by screening a rat brain cDNA library with a cortex-minus-cerebellum subtracted cDNA probe. Conceptual translation of RC3 cDNA sequences indicates that the rat and mouse mRNAs encode identical, novel 78 amino acid proteins. The RC3 protein amino terminus contains a cysteine-rich domain similar to those found in snake venom neurotoxins; the carboxyl terminus contains a collagen-like motif that may function in the assembly of RC3 subunits into a multimeric protein. Western blot experiments with an antiserum to a synthetic peptide corresponding to 27 residues of the 78 residue sequence identify an immunoreactive polypeptide with 18 kDa gel mobility that is likely to correspond to the RC3 protein. Northern blot analysis and in situ hybridization experiments show that RC3 mRNA is highly enriched in rat brain, with restricted expression in neuronal subsets primarily in the cortex, striatum, and hippocampus as well as certain nuclei within the thalamus, hypothalamus, the olfactory bulb.