To be serrate or pinnate: diverse leaf forms of yarrows (Achillea) are linked to differential expression patterns of NAM genes.

Background and Aims: To understand the link between species diversity and phenotype developmental evolution is an important issue in evolutionary biology. Yarrows in the genus Achillea (Asteraceae) show a great diversity in leaf serrate or pinnate dissection patterns. In Arabidopsis thaliana, the development of leaf serration requires the activity of the transcription factor CUC2. Does this regulator also work for leaf dissections of the Asteraceae plants? If so, how do the conserved regulatory 'tools' work differently to produce diverse leaf forms? Methods: Seedling leaf morphology was observed, and morphogenesis of leaf serration or lobes was examined by scanning electron microscopy (SEM). NAM genes, orthologues of arabidopsis CUC2, were isolated from A. acuminata with serrate leaves and A. asiatica with three-pinnatisect leaves, respectively. By means of whole-mount in situ mRNA hybridization and two quantitative gene expression assays, the droplet digital PCR (ddPCR) and quantitative real-time PCR (qPCR), expression patterns of the NAM genes during leaf dissection development were checked in both species for comparison. Key Results: For both species, the development of leaf dissection initiated when a leaf blade was about 300-400 µm long. In A. acuminata, in situ hybridization showed NAM expression signals at leaf margins where teeth are growing, or later on, in the sinuses of the teeth, whilst in A. asiatica, hybridization signals appear not only on leaf margins but further on the margins of leaf lobes. Both ddPCR and qPCR revealed a continuous decline of AacNAM expression from the early to the late developmental stages of a single leaf of A. acuminata, whereas a relatively long maintenance and fluctuation of AasNAM expression was seen in a leaf of A. asiatica. Conclusions: Differential spatiotemporal patterns of NAM expression were found between the two yarrow species during development of leaf dissection. This study provides the first evidence for NAM activity in the development of leaf dissection of the Asteraceae plants, and demonstrates that leaf form diversity is correlated to the altered NAM expression dynamic.

Geographical variation in selection, from phenotypes to molecules.

Molecular technologies now allow researchers to isolate quantitative trait loci (QTLs) and measure patterns of gene sequence variation within chromosomal regions containing important polymorphisms. I develop a simulation model to investigate gene sequence evolution within genomic regions that harbor QTLs. The QTLs influence a trait experiencing geographical variation in selection, which is common in nature and produces obvious differentiation at the phenotypic level. Counter to expectations, the simulations suggest that selection can substantially affect quantitative genetic variation without altering the amount and pattern of molecular variation at sites closely linked to the QTLs. Even with large samples of gene sequences, the likelihood of rejecting neutrality is often low. The exception is situations where strong selection is combined with low migration among demes, conditions that may be common in many plant species. The results have implications for gene sequence surveys and, perhaps more generally, for interpreting the apparently weak connection between levels of molecular and quantitative trait variation within species.

Hybrid origin and differentiation of two tetraploid Achillea species in East Asia: molecular, morphological and ecogeographical evidence.

Achillea (Asteraceae-Anthemideae) offers classical models for speciation by hybridization and polyploidy. Here, we test the suspected allotetraploid origin of two species, Achillea alpina and Achillea wilsoniana between phylogenetically distinct lineages in East Asia. A total of 421 AFLP bands from 169 individuals and 19 populations of five 2x- and two 4x-species were obtained. The data set was analysed with a newly developed model that accounts for polyploidy and assumes lack of recombination between the parental chromosome sets (i.e. disomic inheritance). A. alpina and A. wilsoniana then appear to be allotetraploids between Achillea acuminata-2x (sect. Ptarmica) and Achillea asiatica-2x (sect. Achillea). The two 4x-species share 44% and 48% of their AFLP bands with A. acuminata-2x, and 39% and 38% with A. asiatica-2x, respectively. Eight plastid haplotypes (A-H) were detected by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analyses. A. alpina-4x and A. wilsoniana-4x share haplotype F only with A. asiatica-2x. This is consistent with the hybrid origin(s) involving the latter as the maternal ancestor. This result corroborates our previous DNA sequence data, where A. alpina-4x and A. wilsoniana-4x are also placed close to A. asiatica-2x. Morphology, ecology, and amplified fragment length polymorphism (AFLP) profiles of the two 2x-species are distinct, whereas the two 4x-species, grouped as A. alpina aggregate, form a nearly continuous link between them. Considering all evidence, this 4x-aggregate is regarded as the product of a hybridization between genetically distant 2x-ancestors limited to China and adjacent areas: one A. acuminata-like, and the other A. asiatica-like. The allopolyploid A. alpina agg. exhibits considerable morphological variation and ecological flexibility, and has expanded throughout eastern Asia and to northern North America, far beyond the ranges of their presumed 2x-ancestors.