The Coevolution of Blue-Light Photoreception and Circadian Rhythms

Sunlight is a primary source of energy for life. However, its UV component causes DNA damage. We suggest that the strong UV component of sunlight contributed to the selective pressure for the evolution of the specialized photoreceptor cryptochrome from photolyases involved in DNA repair and propose that early metazoans avoided irradiation by descending in the oceans during the daytime. We suggest further that it is not coincidental that blue-light photoreception evolved in an aquatic environment, since only blue light can penetrate to substantial depths in water. These photoreceptors were then also critical for sensing the decreased luminescence that signals the coming of night and the time to return to the surface. The oceans and the 24-h light-dark cycle therefore provided an optimal setting for an early evolutionary relationship between blue-light photoreception and circadian rhythmicit

Striking circadian neuron diversity and cycling of Drosophila alternative splicing.

Although alternative pre-mRNA splicing (AS) significantly diversifies the neuronal proteome, the extent of AS is still unknown due in part to the large number of diverse cell types in the brain. To address this complexity issue, we used an annotation-free computational method to analyze and compare the AS profiles between small specific groups of Drosophila circadian neurons. The method, the Junction Usage Model (JUM), allows the comprehensive profiling of both known and novel AS events from specific RNA-seq libraries. The results show that many diverse and novel pre-mRNA isoforms are preferentially expressed in one class of clock neuron and also absent from the more standard Drosophila head RNA preparation. These AS events are enriched in potassium channels important for neuronal firing, and there are also cycling isoforms with no detectable underlying transcriptional oscillations. The results suggest massive AS regulation in the brain that is also likely important for circadian regulation

Stabilization and ribosome association of unspliced pre-mRNAs in a yeast upf1- mutant

Nonsense-mediated mRNA decay, the accelerated turnover of mRNAs transcribed from genes containing early nonsense mutations, is dependent on the product of the UPF1 gene in yeast. Mutations that inactivate UPF1 lead to the selective stabilization of mRNAs containing early nonsense mutations but have no effect on the half-lives of almost all other mRNAs. Since the transcripts of nonsense alleles are not typical cellular constituents, we sought to identify those RNAs that comprise normal substrates of the nonsense-mediated mRNA decay pathway. Many yeast pre-mRNAs contain early in-frame nonsense codons and we consider it possible that a role of this pathway is to accelerate the degradation of pre-mRNAs present in the cytoplasm. Consistent with this hypothesis, we find that, in a strain lacking UPF1 function, the CYH2, RP51B, and MER2 pre-mRNAs are stabilized 2- to 5-fold and are associated with ribosomes. We conclude that a major source of early nonsense codon-containing cytoplasmic transcripts in yeast is pre-mRNAs and that the UPF1 protein may be part of a cellular system that ensures that potentially deleterious nonsense fragments of polypeptides do not accumulate

The period gene encodes a predominantly nuclear protein in adult Drosophila

The period gene of Drosophila melanogaster (per) is important for the generation and maintenance of biological rhythms. Previous light microscopic observations indicated that per is expressed in a variety of tissues and cell types and suggested that the per protein (PER) may be present in different subcellular compartments. To understand how PER influences circadian rhythms, it is important to define its subcellular location, especially in adult flies where inducible promoter experiments suggested that it is most relevant to circadian locomotor activity rhythms. To this end, we report the results of an immunoelectron microscopic analysis of wild-type flies and per-beta- galactosidase (beta-gal) fusion gene transgenics using a polyclonal anti-PER antibody or an anti-beta-gal antibody, respectively. Most of the PER antigen and the fusion gene product were located within nuclei, suggesting that PER acts in that subcellular compartment to affect circadian rhythms. The results are discussed in terms of per's possible biochemical functions

Autoreceptor Control of Peptide/Neurotransmitter Corelease from PDF Neurons Determines Allocation of Circadian Activity in Drosophila

SummaryDrosophila melanogaster flies concentrate behavioral activity around dawn and dusk. This organization of daily activity is controlled by central circadian clock neurons, including the lateral-ventral pacemaker neurons (LNvs) that secrete the neuropeptide PDF (pigment dispersing factor). Previous studies have demonstrated the requirement for PDF signaling to PDF receptor (PDFR)-expressing dorsal clock neurons in organizing circadian activity. Although LNvs also express functional PDFR, the role of these autoreceptors has remained enigmatic. Here, we show that (1) PDFR activation in LNvs shifts the balance of circadian activity from evening to morning, similar to behavioral responses to summer-like environmental conditions, and (2) this shift is mediated by stimulation of the Gα,s-cAMP pathway and a consequent change in PDF/neurotransmitter corelease from the LNvs. These results suggest another mechanism for environmental control of the allocation of circadian activity and provide new general insight into the role of neuropeptide autoreceptors in behavioral control circuits

Genome-wide features of neuroendocrine regulation in Drosophila by the basic helix-loop-helix transcription factor DIMMED.

Neuroendocrine (NE) cells use large dense core vesi-cles (LDCVs) to traffic, process, store and secrete neuropeptide hormones through the regulated secre-tory pathway. The dimmed (DIMM) basic helix-loop-helix transcription factor of Drosophila controls the level of regulated secretory activity in NE cells. To pursue its mechanisms, we have performed two in-dependent genome-wide analyses of DIMM’s activi-ties: (i) in vivo chromatin immunoprecipitation (ChIP) to define genomic sites of DIMM occupancy and (ii) deep sequencing of purified DIMM neurons to char-acterize their transcriptional profile. By this com-bined approach, we showed that DIMM binds to con-served E-boxes in enhancers of 212 genes whose expression is enriched in DIMM-expressing NE cells. DIMM binds preferentially to certain E-boxes within first introns of specific gene isoforms. Statistical ma-chine learning revealed that flanking regions of puta-tive DIMM binding sites contribute to its DNA binding specificity. DIMM’s transcriptional repertoire features at least 20 LDCV constituents. In addition, DIMM no-tably targets the pro-secretory transcription factor, creb-A, but significantly, DIMM does not target any neuropeptide genes. DIMM therefore prescribes the scale of secretory activity in NE neurons, by a sys-tematic control of both proximal and distal points in the regulated secretory pathway

The period gene encodes a predominantly nuclear protein in adult Drosophila

The period gene of Drosophila melanogaster (per) is important for the generation and maintenance of biological rhythms. Previous light microscopic observations indicated that per is expressed in a variety of tissues and cell types and suggested that the per protein (PER) may be present in different subcellular compartments. To understand how PER influences circadian rhythms, it is important to define its subcellular location, especially in adult flies where inducible promoter experiments suggested that it is most relevant to circadian locomotor activity rhythms. To this end, we report the results of an immunoelectron microscopic analysis of wild-type flies and per-beta- galactosidase (beta-gal) fusion gene transgenics using a polyclonal anti-PER antibody or an anti-beta-gal antibody, respectively. Most of the PER antigen and the fusion gene product were located within nuclei, suggesting that PER acts in that subcellular compartment to affect circadian rhythms. The results are discussed in terms of per's possible biochemical functions

The Implications of Multiple Circadian Clock Origins

Michael Rosbash considers whether the mechanisms that govern circadian rhythms in different organisms have arisen multiple times in evolution, and discusses the implications for our understanding of circadian clocks

PER-TIM Interactions with the Photoreceptor Cryptochrome Mediate Circadian Temperature Responses in Drosophila

Drosophila cryptochrome (CRY) is a key circadian photoreceptor that interacts with the period and timeless proteins (PER and TIM) in a light-dependent manner. We show here that a heat pulse also mediates this interaction, and heat-induced phase shifts are severely reduced in the cryptochrome loss-of-function mutant cryb. The period mutant perL manifests a comparable CRY dependence and dramatically enhanced temperature sensitivity of biochemical interactions and behavioral phase shifting. Remarkably, CRY is also critical for most of the abnormal temperature compensation of perL flies, because a perL; cryb strain manifests nearly normal temperature compensation. Finally, light and temperature act together to affect rhythms in wild-type flies. The results indicate a role for CRY in circadian temperature as well as light regulation and suggest that these two features of the external 24-h cycle normally act together to dictate circadian phase