Rhododendron agastum represents populations of hybrids between R. irroratum and R. delavayi, which comprise mostly or only F1s, at the two sites examined. The sites differ in that at HDB there was no detected variation in cpDNA type or hybrid class, whereas at ZJY there was variation in both.

Rhododendron (Ericaceae) is a large woody genus in which hybridization may play an important role in evolution and speciation, particularly in the Sino-Himalayan region, where many interfertile species often occur sympatrically. Natural hybridization between Rhododendron delavayi Franch. (= R. arboreum ssp. delavayi) and Rhododendron decorum Franch., which belong to different subsections of subgenus Hymenanthes, was investigated. Material of R. delavayi and R. decorum and their putative hybrids was collected from the wild. On the basis of morphology, chloroplast DNA, nuclear ribosomal DNA, and AFLP profiles, hybrids and parental species were identified. Hybridization occurred in both directions, but was asymmetrical, with R. delavayi as the major maternal parent in the hybrid zone. Most of the hybrids possessed intermediate phenotypes, and amongst the 15 hybrids detected were six F1s, two F2s, one first-generation backcross to R. delavayi, and two first-generation backcrosses to R. decorum. This indicates that, if Rhododendron underwent rapid radiation in this region, it did so in spite of permeable species barriers.

It has been suggested that many plants now found in the arctic originated from ancestors that occurred at high altitudes in the southern mountains of the Northern Hemisphere during the Tertiary. However, this hypothesis has rarely been tested using a molecular phylogenetic approach. Here, we present a fossil‐calibrated molecular phylogeny of Lagotis, an arctic‐alpine genus with the greatest diversity in the Qinghai‐Tibetan Plateau (QTP) and Central Asian mountains, based on five chloroplast (matK, psbA‐trnH, rps16, trnG‐S, trnL‐F) and nuclear ribosomal ITS DNA markers. Within this framework, we infer the ancestral area and biogeographic history of the genus. Four major clades (A–D) within Lagotis were recovered with strong support, which largely correspond to the previous classification of the genus. Within clade A, Lagotis species from QTP were distributed among several subclades, and L. integrifolia from Central Asia was sister to L. glauca and L. minor from the arctic and subarctic region. The Bayesian molecular dating and the ancestral area reconstruction analyses suggested that Lagotis could have originated in the QTP in the Miocene (Tertiary), and that the genus radiated from the Miocene to Pleistocene. The diversification of Lagotis probably took place predominantly in the QTP and it subsequently spread to the Central Asian highlands, followed by northward migration into the arctic. Our results support the hypothesis that the “Central Asiatic Highland Corridor” as an important route for the migration of the flora between the arctic and QTP.

Plant desiccation-related proteins (DRPs) were first identified as pcC13-62 from the resurrection plant Craterostigma plantagineum and it has been suggested they are involved in plant desiccation tolerance. We identified and characterized a plant DRP, which we called MS-desi, in the floral nectar of a subtropical bean species, Mucuna sempervirens (MS). MS-desi is a major nectar protein (nectarin) of the bean plant and expresses exclusively in the stylopodium, where the nectary is located. The full-length MS-desi gene encodes for a protein of 306 amino acids with a molecular mass of 33,248 Da, and possesses a ferritin-like domain and a signal peptide of 30 amino acids. Structural and phylogenetic analysis demonstrated MS-desi has high similarity to members of the plant DRPs, including pcC 13-62 protein. MS-desi has a similar hydropathy profile to that of pcC13-62 with a grand average of hydropathy index of 0.130 for MS-desi and 0.106 for pcC13-62 protein, which is very different from those of dehydrins and late embryogenesis abundant proteins. The protein's secondary structures, both predicted from the amino acid sequence and directly analysed by far UV circular dichroism, showed that MS-desi is mainly composed of alpha helices and is relatively temperature dependent. The structure change is reversible within a wide range of temperatures. Purified MS-desi and raw MS floral nectar showed dose-dependent citrate synthase inhibition activity, but insensitivity to lactate dehydrogenase, suggesting that, unlike dehydrins, it does not act as a chaperone. The overall results constitute, to our knowledge, the first study on a desiccation-related protein in plant floral nectar.

Class II and III chitinases belonging to different glycoside hydrolase families were major nectarins in Rhododendron irroratum floral nectar which showed significant chitinolytic activity. Previous studies have demonstrated antimicrobial activity in plant floral nectar, but the molecular basis for the mechanism is still poorly understood. Two chitinases, class II (Rhchi2) and III (Rhchi3), were characterized from alkaline Rhododendron irroratum nectar by both SDS-PAGE and mass spectrometry. Rhchi2 (27 kDa) and Rhchi3 (29 kDa) are glycoside hydrolases (family 19 and 18) with theoretical pI of 8.19 and 7.04. The expression patterns of Rhchi2 and Rhchi3 were analyzed by semi-quantitative RT-PCR. Rhchi2 is expressed in flowers (corolla nectar pouches) and leaves while Rhchi3 is expressed in flowers. Chitinase in concentrated protein and fresh nectar samples was visualised by SDS-PAGE and chitinolytic activity in fresh nectar was determined spectrophotometrically via chitin-azure. Full length gene sequences were cloned with Tail-PCR and RACE. The amino acid sequence deduced from the coding region for these proteins showed high identity with known chitinases and predicted to be located in extracellular space. Fresh R. irroratum floral nectar showed significant chitinolytic activity. Our results demonstrate that class III chitinase (GH 18 family) also exists in floral nectar. The functional relationship between class II and III chitinases and the role of these pathogenesis-related proteins in antimicrobial activity in nectar is suggested.

No abstract

α-Gal was the most abundant nectarin in selfing CT plants, but was not detected in the nectar of strictly outcrossing sister tobacco species. No function was demonstrated in antimicrobial defence. Therefore, floral nectarins in selfing species maintain their functional significance in reproductive organ development.

Floral nectar can affect the fitness of insect-pollinated plants, through both attraction and manipulation of pollinators. Self-incompatible insect-pollinated plants receive more insect visits than their self-compatible relatives, and the nectar of such species might face increased risk of infestation by pathogens carried by pollinators than self-compatible plants. Proteins in nectar (nectarins) play an important role in protecting the nectar, but little is known regarding nectarins in self-incompatible species. The nectarins from a self-incompatible and insect-pollinated leguminous crop, Canavalia gladiata, were separated using two-dimensional electrophoresis and analysed using mass spectrometry. The predominant nectarin gene was cloned and the gene expression pattern investigated using quantitative real-time PCR. Chitinolytic activity in the nectar was tested with different substrates. The C. gladiata nectar proteome only has one predominant nectarin, an acidic class III chitinase (CaChi3). The full-length CaChi3 gene was cloned, coding for a protein of 298 amino acids with a predicted signal peptide. CaChi3 is very similar to members of the class III chitinase family, whose evolution is dominated by purifying selection. CaChi3 was expressed in both nectary and leaves. CaChi3 has thermostable chitinolytic activity according to glycol-chitin zymography or a fluorogenic substratem but has no lysozyme activity. Chitinase might be a critical protein component in nectar. The extremely simple nectar proteome in C. gladiata disproves the hypothesis that self-incompatible species always have more complex nectar proteomes. Accessibility of nectar might be a significant determinant of the evolutionary pressure to develop nectar defence mechanisms.