Lineage isolation in the face of active gene flow in the coastal plant wild radish is reinforced by differentiated vernalisation responses.

BACKGROUND: The respective role and relative importance of natural selection and gene flow in the process of population divergence has been a central theme in the speciation literature. A previous study presented conclusive evidence that wild radish on Japanese islands comprises two genetically isolated lineages: the southern and northern groups. However, a general understanding of the lineage isolation with frequent seed flow of the coastal plant species is still unclear. We surveyed nucleotide polymorphisms over 14 nuclear loci in 72 individuals across the Japan-Ryukyu Islands Arc to address the demographic history of wild radish utilising the isolation-with-migration (IM) model. In addition, we investigated the flowering times of individuals in different wild radish lineages, with and without cold exposure, to assess their respective vernalisation responses. RESULTS: Coalescent simulations suggested that divergence between the southern and northern lineages of wild radish began ~18,000 years ago, initially during the Last Glacial Maximum (LGM) period. The gene flow from the southern to northern groups was considerably higher than that in the opposite direction, indicating effective dispersal of viable seeds via the northward Kuroshio Current. Our greenhouse experiments indicated that cold exposure was not required for flowering in the southern group, but could advance the date of flowering, suggesting that vernalisation would be facultative in the southern group. In contrast, the northern group was either unable to flower or flowered later without prior cold exposure, and thus had an obligate requirement for cold treatment. CONCLUSIONS: The south-north lineage divergence in wild radish could be triggered by a directional change in the sea current during the ice age, despite gene flow due to the high dispersability and longevity of seeds. We also found that temperature profoundly affected the vernalisation responses of wild radish, which may repress reproductive success and ultimately drive and reinforce intra-specific differentiation between the two lineages of wild radish. This study provides new insights into the maintenance of lineage differentiation with on-going gene flow in coastal plants.

Persistent history of the bird-dispersed arctic-alpine plant Vaccinium vitis-idaea L. (Ericaceae) in Japan.

Arctic-alpine plants have expanded and contracted their ranges in response to the Pleistocene climate oscillations. Today, many arctic-alpine plants have vast distributions in the circumarctic region as well as marginal, isolated occurrences in high mountains at lower latitudes. These marginal populations may represent relict, long-standing populations that have persisted for several cycles of cold and warm climate during the Pleistocene, or recent occurrences that either result from southward step-wise migration during the last glacial period or from recent long-distance dispersal. In light of these hypotheses, we investigated the biogeographic history of the marginal Japanese populations of the widespread arctic-alpine plant Vaccinium vitis-idaea (Ericaceae), which is bird-dispersed, potentially over long distances. We sequenced three nuclear loci and one plastid DNA region in 130 individuals from 65 localities covering its entire geographic range, with a focus on its marginal populations in Japan. We found a homogenous genetic pattern across its enormous range based on the loci analysed, in contrast to the geographically structured variation found in a previous study of amplified fragment length polymorphisms in this species. However, we found several unique haplotypes in the Japanese populations, excluding the possibility that these marginal populations result from recent southward migration. Thus, even though V. vitis-idaea is efficiently dispersed via berries, our study suggests that its isolated populations in Japan have persisted during several cycles of cold and warm climate during the Pleistocene.

Population fragmentation causes randomly fixed genotypes in populations of Arabidopsis kamchatica in the Japanese Archipelago.

Populations of arctic alpine plants likely disappeared and re-colonised several times at the southern edge of their distributions during glacial and interglacial cycles throughout the Quaternary. Range shift and population fragmentation after a glacial period would affect the genetic structure of such plants in southernmost populations. We aimed to elucidate how climatic oscillations influenced the population subsistence of alpine plants in the Japanese Archipelago as one of the southernmost populations, by inferring the genetic structure of Arabidopsis kamchatica subsp. kamchatica and the intraspecific littoral taxon, subsp. kawasakiana. We identified genotypes based on the haplotypes of five nuclear genes and two chloroplast DNA spacers for 164 individuals from 24 populations. Most populations harboured only one private genotype, whereas few polymorphisms were found in each population. Two genetic genealogies were found, suggesting that northern Japanese populations of alpine subsp. kamchatica, subsp. kawasakiana and the northerly subsp. kamchatica in eastern Russia and Alaska clustered and differentiated from populations in central Honshu, western Japan and Taiwan. During climatic oscillations, the genetic structure of extant southernmost populations would have been shaped by strong genetic drift under population fragmentation and randomly fixed to a single genotype among their ancestral polymorphisms.

Development of polymorphic microsatellite loci in the perennial herb Hepatica nobilis var. japonica (Ranunculaceae).

PREMISE OF THE STUDY: Microsatellite markers were developed and characterized in the vulnerable plant Hepatica nobilis var. japonica (Ranunculaceae) to investigate its genetic diversity, population structure, and gene flow. METHODS AND RESULTS: Fourteen microsatellite markers were developed. The number of alleles per locus ranged from one to 12, and the expected heterozygosity per locus ranged from 0.043 to 0.855. Eleven markers were successfully amplified in the cultivar 'Mego' from Japan. CONCLUSIONS: These microsatellite markers can be used to investigate the genetic diversity, population structure, and gene flow of H. nobilis var. japonica.

Distinct Phylogeographic Structures of Wild Radish (Raphanus sativus L. var. raphanistroides Makino) in Japan.

Coastal plants with simple linear distribution ranges along coastlines provide a suitable system for improving our understanding of patterns of intra-specific distributional history and genetic variation. Due to the combination of high seed longevity and high dispersibility of seeds via seawater, we hypothesized that wild radish would poorly represent phylogeographic structure at the local scale. On the other hand, we also hypothesized that wild radish populations might be geographically differentiated, as has been exhibited by their considerable phenotypic variations along the islands of Japan. We conducted nuclear DNA microsatellite loci and chloroplast DNA haplotype analyses for 486 samples and 144 samples, respectively, from 18 populations to investigate the phylogeographic structure of wild radish in Japan. Cluster analysis supported the existence of differential genetic structures between the Ryukyu Islands and mainland Japan populations. A significant strong pattern of isolation by distance and significant evidence of a recent bottleneck were detected. The chloroplast marker analysis resulted in the generation of eight haplotypes, of which two haplotypes (A and B) were broadly distributed in most wild radish populations. High levels of variation in microsatellite loci were identified, whereas cpDNA displayed low levels of genetic diversity within populations. Our results indicate that the Kuroshio Current would have contributed to the sculpting of the phylogeographic structure by shaping genetic gaps between isolated populations. In addition, the Tokara Strait would have created a geographic barrier between the Ryukyu Islands and mainland Japan. Finally, extant habitat disturbances (coastal erosion), migration patterns (linear expansion), and geographic characteristics (small islands and sea currents) have influenced the expansion and historical population dynamics of wild radish. Our study is the first to record the robust phylogeographic structure in wild radish between the Ryukyu Islands and mainland Japan, and might provide new insight into the genetic differentiation of coastal plants across islands.

Benefits of interferon-ß and temozolomide combination therapy for newly diagnosed primary glioblastoma with the unmethylated MGMT promoter: A multicenter study.

BACKGROUND: The aim of the current study was to catalog genomic and epigenomic abnormalities in newly diagnosed glioblastoma patients and determine the correlation among clinical, genetic, and epigenetic profiles and clinical outcome. METHODS: This study retrospectively included 68 consecutive patients who underwent surgical treatment and received standard radiotherapy with temozolomide (TMZ)-based chemotherapy. Of a total of 68 patients, 39 patients (57.4%) received interferon (IFN)-ß in combination of TMZ. RESULTS: The genetic and epigenetic alterations frequently observed were EGFR amplification (51.5%), TP53 mutation (33.8%), CDKN2A loss (32.4%), TP53 loss (16.2%), methylation of the MGMT promoter (33.8%) and IDH1 mutation (5.9%). Multivariate analysis revealed that methylated MGMT promoter and the combination of TMZ and IFN-ß were independent prognostic factors associated with survival. The median survival time (MST) of the patients who received the combination of IFN-ß and TMZ was significantly greater with 19.9 months as compared to the TMZ alone group (12.7 months). Notably, in even patients whose tumors had unmethylated MGMT promoter, the MST prolonged to 17.2 months when receiving TMZ with IFN-ß, compared to 12.5 months in those receiving TMZ without IFN-ß. CONCLUSIONS: Taken together, addition of IFN-ß for newly diagnosed primary GBM achieved a favorable outcome, particularly in patients with unmethylated MGMT promoter.

Expression of tight junction protein claudin-5 in tumor vessels and sinusoidal endothelium in patients with hepatocellular carcinoma.

BACKGROUND: An increase of leaky vasculature is vital for the growth and metastasis of hepatocellular carcinoma (HCC). The paracellular permeability-regulating proteins in tumor vessels and adjacent sinusoids have not been studied in HCC patients. METHODS: Expression of an endothelial tight junction protein claudin-5 (CL-5) and a standard endothelial marker CD34 were immunohistochemically examined in resected specimens from 51 HCC cases. The relationship between hepatitic or fibrotic grade and CL-5 expression pattern in sinusoidal endothelial cells (SECs) was evaluated in the tumor-adjacent tissues. Microvessel density (MVD) highlighted by CD34 or CL-5 was examined in tumor tissues. RESULTS: In the normal liver, a ubiquitous CL-5 expression was seen in SECs, the arteries, and portal veins but not in the central veins. Sinusoidal CL-5 expression was down-regulated according to the increase of hepatitic or fibrotic grade. Poor differentiation and vasculobiliary invasion were significantly associated with a lower CL-5-MVD but not CD34-MVD. By multivariate analysis, vasculobiliary invasion and lower CL-5-MVD were independent factors associated with a lower postoperative overall survival rate. CONCLUSIONS: Attenuated CL-5 expression in SECs may be related to SEC dysfunction in injured liver. Down-regulated CL-5 expression in tumor vessels may serve as a potential marker for poor prognosis in HCC.