Faculty Bibliography

1. Xenopus oocytes injected with rat brain mRNA express a transient K+ current similar to the A current that activates transiently near the threshold for Na+ action potential generation (ISA) seen in somatic recordings from neurons. We used hybrid arrest with antisense oligonucleotides to investigate which of the cloned K+ channel proteins might be components of the channels responsible for the ISA expressed from brain mRNA. An oligonucleotide complementary to a sequence common to all known mammalian Shal-related mRNAs [KV4.1, KV4.2, and KV4.3 (the nomenclature of Sh K+ channel genes of Chandy and colleagues was used in this paper)] blocked the expression of the ISA. An oligonucleotide complementary only to the KV4.2 mRNA, the most abundant Shal-related transcript in rat brain RNA preparations, was also quite efficient in arresting the expression of the ISA from brain. These experiments indicate that Shal-related proteins are important components of the channels carrying the ISA expressed in oocytes injected with brain mRNA. However, there are several significant differences between this ISA and the currents expressed in the same oocytes by in vitro transcribed KV4.1 or KV4.2 cRNA. Most of these differences are eliminated if KV4.1 or KV4.2 cRNA is coinjected with brain poly-(A) RNA treated with antisense oligonucleotides which arrest the expression of the ISA, or with a 2-4Kb rat brain poly-(A) RNA fraction which does not express detectable K+ currents under the same recording conditions. These data support the hypothesis that ISA channels such as those expressed from brain mRNA contain Shal proteins that can be modified by proteins encoded in RNAs that by themselves do not express K+ currents

Homologs of genes mapping to human chromosome 11p15 are located in three distinct, widely separated regions of mouse chromosome 7 (Mmu7). To date, six genes have been localized to the most proximal HSA11p15/Mmu7 homology region, including Ldh3 (encoding lactate dehydrogenase C), Ldh1 (lactate dehydrogenase A), Myod1 (myogenic differentiation factor-1), Tph (tryptophan hydroxylase), Saa1 (serum amyloid-A-1), and Kcnc1 (encoding a Shaw-type voltage-gated potassium channel). To define the overall size and organization of this region of Mmu7, we have established a long-range physical map including the murine Ldh1, Ldh3, Saa, Tph, Kcnc1, and Myod1 genes. Our results demonstrate that these six genes are physically clustered and are distributed throughout a 500-kb interval located just proximal of the pink-eyed dilution (p) locus. These data, together with recent mapping studies within the related region of HSA11p15, demonstrate that gene content and organization within this proximal homology segment have been highly conserved throughout evolution

Human endothelin (ET) A receptor (hETAR) is a G-protein-mediated receptor that binds ET1 with high affinity and ET2 and ET3 with lower affinities. ET1 is the most potent endogenous vasoconstrictor known at this time. When expressed in Xenopus laevis oocytes, hETAR is rapidly desensitized after stimulation with ET1. This desensitization lasts 90-110 min. Human neurokinin A (hNKAR) and human serotonin type 2 receptors were also expressed in the Xenopus system for comparison to hETAR. hNKAR desensitizes for 25-35 min, while the serotonin receptor does not appear to desensitize. To examine the role of the cytoplasmic tail of hETAR in desensitization, deletion mutations were constructed which remove 11, 36, and 51 amino acids from the cytoplasmic tail. The mutations removing 11 and 36 residues were functional, but the mutation removing 51 amino acids was not functional. The two functional mutations have a resensitization time similar to that of hETAR. In summary, the prolonged desensitization time of hETAR is unique for G-protein-mediated receptors and may attenuate the adverse physiological effects of the endothelin family. In addition, the cytoplasmic tail of hETAR does not appear to play a role in desensitization or resensitization of this receptor