Dataset Information

Drosophila TIM binds importin ?1, and acts as an adapter to transport PER to the nucleus.

ABSTRACT: Regulated nuclear entry of clock proteins is a conserved feature of eukaryotic circadian clocks and serves to separate the phase of mRNA activation from mRNA repression in the molecular feedback loop. In Drosophila, nuclear entry of the clock proteins, PERIOD (PER) and TIMELESS (TIM), is tightly controlled, and impairments of this process produce profound behavioral phenotypes. We report here that nuclear entry of PER-TIM in clock cells, and consequently behavioral rhythms, require a specific member of a classic nuclear import pathway, Importin ?1 (IMP?1). In addition to IMP?1, rhythmic behavior and nuclear expression of PER-TIM require a specific nuclear pore protein, Nup153, and Ran-GTPase. IMP?1 can also drive rapid and efficient nuclear expression of TIM and PER in cultured cells, although the effect on PER is mediated by TIM. Mapping of interaction domains between IMP?1 and TIM/PER suggests that TIM is the primary cargo for the importin machinery. This is supported by attenuated interaction of IMP?1 with TIM carrying a mutation previously shown to prevent nuclear entry of TIM and PER. TIM is detected at the nuclear envelope, and computational modeling suggests that it contains HEAT-ARM repeats typically found in karyopherins, consistent with its role as a co-transporter for PER. These findings suggest that although PER is the major timekeeper of the clock, TIM is the primary target of nuclear import mechanisms. Thus, the circadian clock uses specific components of the importin pathway with a novel twist in that TIM serves a karyopherin-like role for PER.

SUBMITTER: Jang AR

PROVIDER: S-EPMC4335507 | biostudies-literature | 2015 Feb

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Drosophila TIM binds importin α1, and acts as an adapter to transport PER to the nucleus.

Jang A Reum AR Moravcevic Katarina K Saez Lino L Young Michael W MW Sehgal Amita A

PLoS genetics 20150212 2

Regulated nuclear entry of clock proteins is a conserved feature of eukaryotic circadian clocks and serves to separate the phase of mRNA activation from mRNA repression in the molecular feedback loop. In Drosophila, nuclear entry of the clock proteins, PERIOD (PER) and TIMELESS (TIM), is tightly controlled, and impairments of this process produce profound behavioral phenotypes. We report here that nuclear entry of PER-TIM in clock cells, and consequently behavioral rhythms, require a specific me ...[more]

PMID: 25674790

Similar Datasets

Project description:Bovine ephemeral fever virus (BEFV) is an arthropod-borne rhabdovirus that is classified as the type species of the genus Ephemerovirus. In addition to the five canonical rhabdovirus structural proteins (N, P, M, G, and L), the large and complex BEFV genome contains several open reading frames (ORFs) between the G and L genes (α1, α2/α3, β, and γ) encoding proteins of unknown function. We show that the 10.5-kDa BEFV α1 protein is expressed in infected cells and, consistent with previous predictions based on its structure, has the properties of a viroporin. Expression of a BEFV α1-maltose binding protein (MBP) fusion protein in Escherichia coli was observed to inhibit cell growth and increase membrane permeability to hygromycin B. Increased membrane permeability was also observed in BEFV-infected mammalian cells (but not cells infected with an α1-deficient BEFV strain) and in cells expressing a BEFV α1-green fluorescent protein (GFP) fusion protein, which was shown by confocal microscopy to localize to the Golgi complex. Furthermore, the predicted C-terminal cytoplasmic domain of α1, which contains a strong nuclear localization signal (NLS), was translocated to the nucleus when expressed independently, and in an affinity chromatography assay employing a GFP trap, the full-length α1 was observed to interact specifically with importin β1 and importin 7 but not with importin α3. These data suggest that, in addition to its function as a viroporin, BEFV α1 may modulate components of nuclear trafficking pathways, but the specific role thereof remains unclear. Although rhabdovirus accessory genes occur commonly among arthropod-borne rhabdoviruses, little is known of their functions. Here, we demonstrate that the BEFV α1 ORF encodes a protein which has the structural and functional characteristics of a viroporin. We show that α1 localizes in the Golgi complex and increases cellular permeability. We also show that BEFV α1 binds importin β1 and importin 7, suggesting that it may have a yet unknown role in modulating nuclear trafficking. This is the first functional analysis of an ephemerovirus accessory protein and of a rhabdovirus viroporin.

Project description:Circadian (approximately 24-h) rhythms are governed by endogenous biochemical oscillators (clocks) that in a wide variety of organisms can be phase shifted (i.e., delayed or advanced) by brief exposure to light and changes in temperature. However, how changes in temperature reset circadian timekeeping mechanisms is not known. To begin to address this issue, we measured the effects of short-duration heat pulses on the protein and mRNA products from the Drosophila circadian clock genes period (per) and timeless (tim). Heat pulses at all times in a daily cycle elicited dramatic and rapid decreases in the levels of PER and TIM proteins. PER is sensitive to heat but not light, indicating that individual clock components can markedly differ in sensitivity to environmental stimuli. A similar resetting mechanism involving delays in the per-tim transcriptional-translational feedback loop likely underlies the observation that when heat and light signals are administered in the early night, they both evoke phase delays in behavioral rhythms. However, whereas previous studies showed that the light-induced degradation of TIM in the late night is accompanied by stable phase advances in the temporal regulation of the PER and TIM biochemical rhythms, the heat-induced degradation of PER and TIM at these times in a daily cycle results in little, if any, long-term perturbation in the cycles of these clock proteins. Rather, the initial heat-induced degradation of PER and TIM in the late night is followed by a transient and rapid increase in the speed of the PER-TIM temporal program. The net effect of these heat-induced changes results in an oscillatory mechanism with a steady-state phase similar to that of the unperturbed control situation. These findings can account for the lack of apparent steady-state shifts in Drosophila behavioral rhythms by heat pulses applied in the late night and strongly suggest that stimulus-induced changes in the speed of circadian clocks can contribute to phase-shifting responses.

Dataset Information

Drosophila TIM binds importin ?1, and acts as an adapter to transport PER to the nucleus.

Publications

Drosophila TIM binds importin α1, and acts as an adapter to transport PER to the nucleus.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets