In this study we have isolated and sequenced human cDNAs for the phosphoproteins DARPP-32, ARPP-21, and ARPP-16/19, and have compared these sequences to previously characterized bovine and rat cDNAs. In situ hybridization and Northern blot analysis with the human cDNA probes were used to study the expression of mRNAs encoding ARPP-16/19, ARPP-21, and DARPP-32 in human postmortem brain tissue. In situ hybridization was performed using horizontal whole hemisphere sections. Five representative levels of the brain ranging from 71 mm to 104 mm ventral to vertex were examined. All three probes showed distinct hybridization patterns in the caudate nucleus, putamen, nucleus accumbens, and the amygdaloid complex. For ARPP-16/19 mRNA, a hybridization signal comparable to the signal in caudate nucleus, putamen, and nucleus accumbens was also detected in the neocortex. ARPP- 21 and DARPP-32 mRNA, on the other hand, were present in lower levels in neocortical regions. DARPP-32 mRNA was abundant in the cerebellar cortex at the level of the Purkinje cell layer. High levels of ARPP- 16/19 and ARPP-21 mRNA were also found in the cerebellar cortex, where they were confined to deeper layers. The present result demonstrate that mRNAs for the three phosphoproteins are expressed in overlapping, but also distinct, areas of the human brain that in many cases coincide with previously described distribution of the dopamine D1 receptor.

A cDNA clone for the mRNA of bovine ARPP-21 (cAMP-regulated phosphoprotein, Mr = 21,000 as determined by SDS-PAGE) was isolated from a modified Okayama-Berg plasmid library. Transformed Escherichia coli colonies were screened by in situ colony hybridization with 2 different oligonucleotide probes derived from the amino acid sequence of the bovine protein. Sequence analysis of the longest cDNA clone, pTKAI [2407 nucleotides plus a poly(A) tail], revealed a 267-nucleotide-long coding region in agreement with the bovine ARPP-21 amino acid sequence (Williams et al., 1989). Southern blot analysis of total bovine genomic DNA raised the possibility that there may be 2 genes coding for ARPP-21. Northern blot analysis of total cellular RNA from bovine caudate nucleus and other brain regions demonstrated the existence of 2 major mRNA species, 2.5 and 1.0 kb in length, probably derived from use of alternate polyadenylation sites. There was a differential expression of these 2 mRNAs within the brain. Both ARPP-21 mRNAs were most abundant in the caudate nucleus, where the concentration of the protein is highly enriched.

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