Cloning of a cDNA Encoding the Rat High Molecular Weight Neurofilament Peptide (NF-H): Developmental and Tissue Expression in the Rat, and Mapping of Its Human Homologue to Chromosomes 1 and 22

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 86; no. 7; pp. 2463 - 2467
• Main Authors: Lieberburg, Ivan, Spinner, Nancy, Synder, Susan, Anderson, John, Goldgaber, Dmitri, Smulowitz, Mildred, Carroll, Zev, Emanuel, Beverly, Breitner, John, Rubin, Lee
• Format: Journal Article
• Language: English
• Published: Washington, DC National Academy of Sciences of the United States of America 01.04.1989; National Acad Sciences
• Subjects: 550401 - Genetics- Tracer Techniques; Aging; Amino Acid Sequence; Amino acids; ANIMAL CELLS; ANIMALS; Base Sequence; BASIC BIOLOGICAL SCIENCES; BETA DECAY RADIOISOTOPES; BETA-MINUS DECAY RADIOISOTOPES; Biological and medical sciences; BODY; BRAIN; Brain - embryology; Brain - growth & development; CENTRAL NERVOUS SYSTEM; Chromosome Mapping; CHROMOSOMES; Chromosomes, Human, Pair 1; Chromosomes, Human, Pair 22; CLONING; Cloning, Molecular; Complementary DNA; DAYS LIVING RADIOISOTOPES; DISTRIBUTION; DNA; DNA - genetics; DNA HYBRIDIZATION; DNA-CLONING; Female; Fundamental and applied biological sciences. Psychology; Generally accepted auditing standards; Genes; Genes. Genome; GENETIC MAPPING; HETEROCHROMOSOMES; Humans; HYBRIDIZATION; Intermediate Filament Proteins - biosynthesis; Intermediate Filament Proteins - genetics; ISOTOPES; LIGHT NUCLEI; MAMMALS; MAN; MAPPING; MESSENGER-RNA; Molecular and cellular biology; Molecular genetics; Molecular Sequence Data; MOLECULAR WEIGHT; NERVE CELLS; Nerve tissue; NERVOUS SYSTEM; Neurofilament Proteins; Neurons; NUCLEI; NUCLEIC ACIDS; NUCLEOTIDES; ODD-ODD NUCLEI; ORGANIC COMPOUNDS; ORGANS; PEPTIDES; PHOSPHORUS 32; PHOSPHORUS ISOTOPES; Placenta - metabolism; Pregnancy; PRIMATES; PROTEINS; RADIOISOTOPES; RATS; RECOMBINANT DNA; RNA; RODENTS; Sequence Homology, Nucleic Acid; SOMATIC CELLS; TISSUE DISTRIBUTION; TRANSCRIPTION; VERTEBRATES; Vertebrata; Neurofilament; Mammalia; Complementary DNA; Peptides; Rat; Rodentia; Development; Chromosome; Homology; Molecular cloning; Gene expression
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.86.7.2463

Neurofilaments (NFs) are the intermediate filaments specific to nervous tissue. They are probably essential to the tensile strength of the neuron, as well as to transport of molecules and organelles within the axon. Three peptides with apparent molecular masses of approximately 68 (NF-L), 145 (NF-M), and 200 (NF-H) kDa appear to be the major components of NF. The expression of these peptides is specific to nervous tissue and is developmentally regulated. Recently, complete cDNAs encoding NF-L and NF-M, and partial cDNAs encoding NF-H, have been described. To better understand the normal and pathophysiology of NFs we chose to clone the cDNA encoding the rat NF-H peptide. Using monoclonal antibodies that recognized NF-H, we screened a rat brain λ gt11 library and identified a clone that contained a 2100-nucleotide cDNA insert representing the carboxyl-terminal portion of the NF-H protein. Anti-fusion protein antibodies recognized the NF-H peptide on immunoblots and stained fibrillar structures only in neurons. The cDNA recognized a 4500-nucleotide polyadenylylated mRNA that was present only in nervous tissue and a 3500-nucleotide mRNA in adrenal. Brain NF-H mRNA levels were tightly developmentally regulated and paralleled the levels of NF-H peptide on immunoblots. Nuclear runoff studies showed that the 20-fold developmental increase in the NF-H message was due only in part to a 4-fold increase in its transcription rate. Levels of NF-H mRNA varied 20-fold among brain regions, with highest levels in pons/medulla, spinal cord, and cerebellum, and lowest levels in olfactory bulb and hypothalamus. Transcription studies revealed only a 2-fold difference in the transcription rates among these brain regions. Based on these results, we infer that half of the developmental increase and most of the interregional variation in the levels of the NF-H mRNA are mediated through message stabilization. Sequence information revealed that the carboxyl-terminal region of the NF-H peptide contained a unique serine-, proline-, alanine-, glutamic acid-, and lysine-rich repeat. The serine residues are likely sites of phosphorylation in the mature peptide. Genomic blots revealed a single copy of the gene in the rat genome and two copies in the human genome. In situ hybridizations performed on human chromosomes mapped the NF-H gene to chromosomes 1 and 22. Whether one copy is a pseudogene remains to be determined.