cDNA cloning and nuclear localization of the circadian neuropeptide designated as pigment-dispersing factor PDF in the cricket Gryllus bimaculatus.
Pigment-dispersing factor (PDF) was recently reported to be a principal circadian neuromodulator involved in transmitting circadian rhythms of daily locomotion in insects. In Drosophila, PDF functions in some of the neurons expressing the clock genes period, timeless, Clock, and cycle, and those clock genes in turn regulate pdf gene expression. In the present study, we cloned a cDNA encoding PDF in the brain of a nocturnal insect, the cricket Gryllus bimaculatus, and found that an isolated clone (310 bp) codes for an extraordinarily short precursor protein with no definite signal sequence, but a nuclear localization signal (NLS)-like sequence instead. The cricket PDF exhibits high sequence identity (78-94%) and similarity (89-100%) to insect PDFs and also to crustacean beta-PDH peptides. In the optic lobes of G. bimaculatus there are PDF-immunoreactive neurons in both the medulla and lamina neuropiles. Among the strongly immunoreactive lamina PDF neurons, on electron microscopy we identified cells exhibiting distinct staining that is not only cytoplasmic but also nuclear. When GFP-fused PDF precursor proteins were expressed in COS-7 cells, distinct translocation of the fusion protein into the nucleus was observed. This is the first finding of PDF peptide in the nucleus, which suggests a fundamental role of PDF peptide per se in the circadian clock system.
Chuman
Yoshiro
Y
Laboratory of Structure-Function Biochemistry, Department of Chemistry, Faculty and Graduate School of Sciences, Kyushu University, Fukuoka 812-8581, Japan.
Matsushima
Ayami
A
Sato
Seiji
S
Tomioka
Kenji
K
Tominaga
Yoshiya
Y
Meinertzhagen
Ian A
IA
Shimohigashi
Yasuyuki
Y
Shimohigashi
Miki
M
D021581:Active Transport, Cell Nucleus; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002467:Cell Nucleus / Q000378:metabolism*; D002940:Circadian Rhythm / Q000235:genetics*; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000032:analysis; D029721:Drosophila Proteins; D049452:Green Fluorescent Proteins; D006135:Gryllidae; D008164:Luminescent Proteins / Q000378:metabolism; D008969:Molecular Sequence Data; D009479:Neuropeptides / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D021382:Protein Sorting Signals / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D020539:Sequence Analysis, Protein; D017386:Sequence Homology, Amino Acid
D018076:DNA, Complementary; D029721:Drosophila Proteins; D008164:Luminescent Proteins; D009479:Neuropeptides; D021382:Protein Sorting Signals; D011993:Recombinant Fusion Proteins; C113883:pdf protein, Drosophila; D049452:Green Fluorescent Proteins