BIOLOGY OF THE MALE REPRODUCTIVE TRACT
7:30 AM-10:00 AM
(580) DETECTION OF POTENTIAL REGULATORS OF NITRERGIC NEUROTRANSMISSION OF PENILE ERECTION IN THE RAT PENIS.
Qian, Ansha1, Rajfer, Jacob1, Gonzalez-Cadavid, Nestor1, 1 Department of Urology, UCLA School of Medicine, Harbor-UCLA REI, Torrance, CA
ABSTRACT- Penile erection is controlled by the activity of neuronal nitric oxide synthase (nNOS) variants to produce nitric oxide (NO) during nitrergic neurotransmission in the hypothalamus, spinal cord, and penis. nNOS activity is modulated by binding of Ca2+ and certain regulatory proteins (PIN, PSD95, NMDA receptor (NMDAR)), which also affect nNOS association with the cytoskeleton. Both PIN, a light chain dynein (LCD), and a variant of the NMDAR are expressed in the penis but their role in modulating nNOS is unclear. We hypothesized that protein interactions with the PDZ domain of nNOS involve additional cytoskeleton and Ca2+ channel-linked proteins, and that changes in their expression may lead to a reduced NO synthesis as is seen in aging-related erectile dysfunction. To examine these questions, we screened peptide interactions with rat recombinant nNOS in an E.coli-based dodecapeptide display library. Some of the plasmid clones (40) for the bound peptides, were amplified and sequenced. Peptide sequence homologies with databanks were selected based on neural tissue expression, potential role in the nitrerrgic cascade, and expression from at least 4 clones. In order of clone abundance, they include: cytoplasmic dynein heavy chain 2 (CDHC2), neuronal myosin heavy chain (NMHC), glutamate receptor (GR), voltage-dependent Ca2+ channel, NMDAR, purine receptor (PZY), PSD95, PDZ proteins, and calmodulin binding protein. RNA from penile shaft tissue from young (5 month old) and old (24 month old) rats was analyzed by an Affymetrix Genechip (8,000 genes). All putatively nNOS interacting proteins were expressed in the penile tissue, and a few were among the 139 and 117 genes up- and down-regulated (>2-fold), respectively, by aging. We postulate that some of the identified interacting proteins modulate PIN inhibition, or nNOS binding to cytoskeleton and its association with Ca2+ channel-related receptors. These novel interactions for nNOs in penile tissue may have relevance to the defective NO synthesis in erectile dysfunction.
KEY WORDS: penile erection, nitric oxide, microarray, peptide display