Induced nitric oxide synthesis is dependent on induced alternatively spliced CAT-2 encoding L-arginine transport in brain astrocytes.
The inducible isoform II of nitric-oxide synthase (iNOS) was recently cloned from brain and identified in astroglial cells. Induced nitric oxide biosynthesis occurs in brain cells only if extracellular cerebrospinal fluid contains -arginine. This study demonstrates for the first time that induced iNOS activity is strictly dependent on concomitant induction of an alternatively spliced transcript of the cat-2 gene encoding high affinity -arginine transporter System y+ in cultured rat astrocytes. Inhibition profiles of radiolabeled -arginine and -leucine uptake identified the dominance of Na+-independent transport System y+ serving cationic amino acids, with insignificant activities of Systems y+L, bo,+, or Bo,+. A reverse transcription-polymerase chain reaction/sequencing/cloning strategy was used to identify a single 123-base nucleotide sequence coding the high affinity domain of alternatively spliced CAT-2 (not CAT-2a) in astrocytes activated by lipopolysaccharide/interferon-gamma. Using this sequence as a cDNA probe, it was determined that CAT-2 mRNA, iNOS mRNA, and System y+ activity were concomitantly and strongly induced in astrocytes. Constitutive CAT-1 mRNA was weakly present in neurons and astrocytes, was not inducible in either cell type, and contributed
Alternative Splicing, Amino Acid Sequence, Animals, Arginine, Astrocytes, Base Sequence, Biological Transport, Brain, Carrier Proteins, Cell Membrane, Enzyme Induction, Gene Expression, Interferon-gamma, Lipopolysaccharides, Membrane Glycoproteins, Membrane Proteins, Molecular Sequence Data, Nitric Oxide, Nitric Oxide Synthase, RNA, Messenger, Rats, Rats, Inbred WKY, Receptors, Virus, Recombinant Proteins
Medical Subject Headings
Alternative Splicing; Amino Acid Sequence; Animals; Arginine; Astrocytes; Base Sequence; Biological Transport; Brain; Carrier Proteins; Cell Membrane; Enzyme Induction; Gene Expression; Interferon-gamma; Lipopolysaccharides; Membrane Glycoproteins; Membrane Proteins; Molecular Sequence Data; Nitric Oxide; Nitric Oxide Synthase; RNA, Messenger; Rats; Rats, Inbred WKY; Receptors, Virus; Recombinant Proteins
The Journal of biological chemistry
Digital Object Identifier (DOI)
Stevens, B R; Kakuda, D K; Yu, K; Waters, M; Vo, C B; and Raizada, M K, "Induced nitric oxide synthesis is dependent on induced alternatively spliced CAT-2 encoding L-arginine transport in brain astrocytes." (1996). Translational Neuroscience. 1495.