Identification of transcriptional regulators of neuropeptide FF gene expression.
Neuropeptide FF (NPFF) is an RF-amide peptide with pleiotropic functions in the mammalian central nervous system, including pain modulation, opiate interactions, cardiovascular regulation and neuroendocrine effects. To gain insights into the transcriptional mechanisms that regulate NPFF gene expression, we cloned and sequenced 9.8 and 1.5 kb of the mouse and rat NPFF 5'-flanking region, respectively. Regions with high sequence homology between mouse, rat and human were expected to have high probability to interact with regulatory proteins and were studied further. Electromobility shift assays revealed one region that may interact with the homeobox proteins Oct-1, PDX1, Pit-1 and MEIS and two consensus DRE sites that bind a nuclear protein, which was identified as the downstream regulatory element antagonistic modulator DREAM by supershift assays. The distribution of NPFF gene expression was examined in the mouse using in situ hybridization and RT-PCR. NPFF expression was also evident during mouse embryogenesis. A fixed transcription initiation site for the mouse NPFF gene was found. A novel splice variant with a retained intron of the NPFF gene was characterized. Chimeric luciferase reporter gene constructs for the mouse NPFF gene revealed a minimal promoter region and a region with transcriptional suppressor features. An NGF responsive area was found using mouse NPFF reporter gene constructs. We postulate that Oct-1, PDX1, Pit-1, MEIS and DREAM are likely transcriptional regulators of NPFF gene expression.
Nystedt
Johanna M
JM
Department of Biology, Abo Akademi University, Biocity 2. floor, Tykistökatu 6 A, 20520 Turku, Finland. johanna.nystedt@veripalvelu.fi
Brandt
Annika
A
Vilim
Ferdinand S
FS
Ziff
Edward B
EB
Panula
Pertti
P
D024506:5' Flanking Region / Q000235:genetics; D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002490:Central Nervous System / Q000378:metabolism; D002874:Chromosome Mapping / Q000662:veterinary; D003001:Cloning, Molecular; D024202:Electrophoretic Mobility Shift Assay; D005786:Gene Expression Regulation; D018398:Homeodomain Proteins / Q000502:physiology; D006801:Humans; D051605:Kv Channel-Interacting Proteins / Q000235:genetics; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008969:Molecular Sequence Data; D000075064:Myeloid Ecotropic Viral Integration Site 1 Protein; D009363:Neoplasm Proteins / Q000502:physiology; D050811:Octamer Transcription Factor-1 / Q000502:physiology; D009842:Oligopeptides / Q000235:genetics*; D051381:Rats; D016415:Sequence Alignment; D015534:Trans-Activators / Q000502:physiology; D050817:Transcription Factor Pit-1 / Q000502:physiology; D014157:Transcription Factors / Q000235:genetics*
D018398:Homeodomain Proteins; D051605:Kv Channel-Interacting Proteins; D000075064:Myeloid Ecotropic Viral Integration Site 1 Protein; D009363:Neoplasm Proteins; D050811:Octamer Transcription Factor-1; D009842:Oligopeptides; C494166:Pou2f1 protein, mouse; D015534:Trans-Activators; D050817:Transcription Factor Pit-1; D014157:Transcription Factors; C084010:pancreatic and duodenal homeobox 1 protein; C053651:phenylalanyl-leucyl-phenylalanyl-glutaminyl-prolyl-glutaminyl-arginyl-phenylalaninamide