No abstract

NELF, a protein identified in migratory GnRH neurons, is predominantly nuclear and alternatively spliced. However, specific NELF splice variants expressed in immortalized GnRH neuronal cell lines from mouse and human are not known. RNA from migratory (GN11 and NLT) and postmigratory (GT1-7) cells in mouse, and (FNCB4-hTERT) cells in human was subjected to RT-PCR. RT-PCR products were cloned, electrophoresed on denaturing gradient gels and sequenced. In addition, quantitative RT-PCR was performed using variant-specific primers. Western blot and immunofluorescence using confocal microscopy were performed for selected variants. Nelf variant 2 (v2), which contains a nuclear localization signal (NLS), was the predominant variant in all mouse and human GnRH neurons. Variants without a NLS (v3 in mouse; v4 in human) were identified. In mouse, v2 protein expression was nuclear, while v3 was non-nuclear. In mouse GnRH neurons, six Nelf splice variant transcripts were identified, including three previously unreported variants. In human, four NELF variant transcripts were observed. In both mouse and human, nuclear and non-nuclear variant transcript and protein were identified, explaining variable NELF cellular localization.

Conservation of the gene pool of F. pennsylvanica (green ash), the species most susceptible to emerald ash borer (EAB), requires a range-wide genetic assessment of the population dynamics to guide seed collection efforts. An assessment of gene flow into natural stands from ubiquitously distributed green ash and F. americana (white ash) cultivars is also essential. We used 16 EST-SSR markers to genotype 1291 trees from 48 naturally regenerated populations of green ash, 19 green ash cultivars and 10 white ash from a Fraxinus species collection. We did not find evidence for latitudinal gradients in allele richness or higher differentiation in northern range edge populations. Analysis of population substructure revealed two major groups of green ash, one dominant in northwestern locations, one dominant in southern locations and extensive admixture in midwestern and eastern locations. We identified a third group as white ash, including 140 individuals with interspecific admixture in eastern and southern populations. We detected cultivar parentage with high confidence in 172 individuals in 34 of the 48 populations. We conclude that high dispersal capacity, broad adaptability, and gene flow from many cultivars into predominately northern regions, plays a role in the comparatively low regional population substructure in the Northwest. The high frequency of cultivar propagule dispersal and extent of interspecific admixture detected in our study suggests that under rapidly changing conditions, local populations may or may not be locally adapted and standing genetic variation may not be of long standing.

The emerald ash borer (Agrilus planipennis, EAB) invasion in North America threatens most North American Fraxinus species, including green ash (F. pennsylvanica), the mostly widely distributed species. A small number of green ash (lingering ash, 0.1-1%) survive years of heavy EAB attack and kill more EAB larvae when challenged in greenhouse studies than susceptible controls. We combined untargeted metabolomics with intensive phenotyping of segregating F1 progeny from susceptible or lingering ash parents to detect chemotypes associated with defensive responses to EAB. We examined three contrasting groups: low larval kill (0-25% of larvae killed), high larval kill (55-95% of larvae killed) and uninfested. Contrasting the chemotypes of these groups revealed evidence of an induced response to EAB. Infested trees deployed significantly higher levels of select secoiridoids than uninfested trees. Within the infested group, the low larval kill (LLK) individuals deployed significantly higher levels of select secoiridoids than the high larval kill (HLK) individuals. The HLK individuals deployed significantly higher concentrations of three metabolites annotated as aromatic alkaloids compared to the LLK and uninfested individuals. We propose a two-part model for the North American Fraxinus response to EAB wherein every individual has the capacity to detect and respond to EAB, but only certain trees mount an effective defense, killing enough EAB larvae to prevent or minimize lethal damage to the vascular system. Integration of intensive phenotyping of structured populations with metabolomics reveals the multi-faceted nature of the defenses deployed in naive host populations against invasive species.