The S-Phase Cyclin Clb5 Promotes rRNA Gene (rDNA) Stability by Maintaining Replication Initiation Efficiency in rDNA.

Regulation of replication origins is important for complete duplication of the genome, but the effect of origin activation on the cellular response to replication stress is poorly understood. The budding yeast rRNA gene (rDNA) forms tandem repeats and undergoes replication fork arrest at the replication fork barrier (RFB), inducing DNA double-strand breaks (DSBs) and genome instability accompanied by copy number alterations. Here, we demonstrate that the S-phase cyclin Clb5 promotes rDNA stability. Absence of Clb5 led to reduced efficiency of replication initiation in rDNA but had little effect on the number of replication forks arrested at the RFB, suggesting that arrival of the converging fork is delayed and forks are more stably arrested at the RFB. Deletion of CLB5 affected neither DSB formation nor its repair at the RFB but led to homologous recombination-dependent rDNA instability. Therefore, arrested forks at the RFB may be subject to DSB-independent, recombination-dependent rDNA instability. The rDNA instability in clb5Δ was not completely suppressed by the absence of Fob1, which is responsible for fork arrest at the RFB. Thus, Clb5 establishes the proper interval for active replication origins and shortens the travel distance for DNA polymerases, which may reduce Fob1-independent DNA damage.

Changes in a Cone Opsin Repertoire Affect Color-Dependent Social Behavior in Medaka but Not Behavioral Photosensitivity.

Common ancestors of vertebrates had four types of cone opsins: short-wavelength sensitive 1 (SWS1), SWS2, rhodopsin 2 (RH2), and long-wavelength sensitive (LWS) types. Whereas fish and birds retain all the types, mammals have lost two of them (SWS2 and RH2) possibly because of their nocturnal lifestyle during the Mesozoic Era. Considering that the loss of cone opsin types causes so-called color blindness in humans (e.g., protanopia), the ability to discriminate color by trichromatic humans could be lower than that in potentially tetrachromatic birds and fish. Behavioral studies using color-blind (cone opsin-knockout) animals would be helpful to address such questions, but it is only recently that the genome-editing technologies have opened up this pathway. Using medaka as a model, we introduced frameshift mutations in SWS2 (SWS2a and/or SWS2b) after detailed characterization of the loci in silico, which unveiled the existence of a GC-AG intron and non-optic expressed-sequence-tags (ESTs) that include SWS2a in part. Transcripts from the mutated SWS2 loci are commonly reduced, suggesting that the SWS2a/b-double mutants could produce, if any, severely truncated (likely dysfunctional) SWS2s in small amounts. The mutants exhibited weakened body color preferences during mate choice. However, the optomotor response (OMR) test under monochromatic light revealed that the mutants had no defect in spectral sensitivity, even at the absorbance maxima (λmax) of SWS2s. Evolutionary diversification of cone opsins has often been discussed in relation to adaptation to dominating light in habitats (i.e., changes in the repertoire or λmax are for increasing sensitivity to the dominating light). However, the present results seem to provide empirical evidence showing that acquiring or losing a type of cone opsin (or changes in λmax) need not substantially affect photopic or mesopic sensitivity. Other points of view, such as color discrimination of species-specific mates/preys/predators against habitat-specific backgrounds, may be necessary to understand why cone opsin repertories are so various among animals.

Evolutionary history of the medaka long-wavelength sensitive genes and effects of artificial regression by gene loss on behavioural photosensitivity.

Tandem gene duplication has led to an expansion of cone-opsin repertoires in many fish, but the resulting functional advantages have only been conjectured without empirical demonstration. Medaka (Oryzias latipes and O. sakaizumii) have eight (two red, three green, two blue, and one violet) cone opsin genes. Absorbance maxima (λmax) of the proteins vary from 356 nm to 562 nm, but those of the red opsins (long-wavelength sensitive; LWS) are nearly identical, obscuring the necessity of their coexistence. Here, we compared the LWSa and LWSb loci of these sister species and found that the gene duplication occurred long before the latipes-sakaizumii speciation (4-18 million years ago), and the high sequence similarity between the paralogues is the result of at least two events of gene conversion. These repetitive gene conversions would indicate the importance for medaka of retaining two identical LWSs in the genome. However, a newly established medaka mutant with a single LWS showed no defect in LWS expression or behavioural red-light sensitivity, demonstrating functional redundancy of the paralogs. Thus, as with many other genes after whole-genome duplication, the redundant LWS might be on the way to being lost from the current cone opsin repertoire. Thus, non-allelic gene conversion may temporarily provide an easier and more frequent solution than gene loss for reducing genetic diversity, which should be considered when assessing history of gene evolution by phylogenetic analyses.

Results of second-stage screening for skin cancers in Oita Prefecture, Japan.

We performed skin cancer screenings for 2 or 3 days annually from 2006 through 2013 in Oita Prefecture, Japan. Screening of approximately 3000 people in total allowed us to identify and treat several skin cancers, including five cases of malignant melanoma, four of squamous cell carcinoma, 16 of basal cell carcinoma, 11 of Bowen's disease, 17 of actinic keratosis, one of extramammary Paget's disease and one of metastatic breast carcinoma. The sensitivity and specificity for the category defined by an identified lesion associated with risk of cancer and requiring further examination (category C) were 92.7% and 95%, respectively. We cannot estimate the outcome of our skin cancer screenings in terms of cancer mortality because of the small number of subjects examined and the brief follow-up period. However, we did estimate the effectiveness of these screenings in terms of stages or sizes of cancerous lesions. The relative numbers of subjects with malignant melanoma at various clinical stages, identified during skin cancer screenings and during a routine visit to our hospital, were significantly different. We also compared, statistically, the sizes of lesions in Bowen's disease that were found during cancer screenings and during a direct visit to our hospital. The former lesions were smaller than the latter. Our data suggest the benefits of our skin cancer screenings and the importance of campaigns and education to encourage people to visit dermatologists for the detection of skin cancers at an early stage.