Phylogeny of Impatiens (Balsaminaceae): integrating molecular and morphological evidence into a new classification

Impatiens L. is one of the largest angiosperm genera, containing over 1000 species, and is notorious for its taxonomic difficulty. Here, we present, to our knowledge, the most comprehensive phylogenetic analysis of the genus to date based on a total evidence approach. Forty‐six morphological characters, mainly obtained from our own investigations, are combined with sequence data from three genetic regions, including nuclear ribosomal ITS and plastid atpB‐rbcL and trnL‐F. We include 150 Impatiens species representing all clades recovered by previous phylogenetic analyses as well as three outgroups. Maximum‐parsimony and Bayesian inference methods were used to infer phylogenetic relationships. Our analyses concur with previous studies, but in most cases provide stronger support. Impatiens splits into two major clades. For the first time, we report that species with three‐colpate pollen and four carpels form a monophyletic group (clade I). Within clade II, seven well‐supported subclades are recognized. Within this phylogenetic framework, character evolution is reconstructed, and diagnostic morphological characters for different clades and subclades are identified and discussed. Based on both morphological and molecular evidence, a new classification outline is presented, in which Impatiens is divided into two subgenera, subgen. Clavicarpa and subgen. Impatiens; the latter is further subdivided into seven sections.     

Analyzing the feasibility of discriminating between collagen types I and II using polarization‐resolved second harmonic generation

According to previous studies, the nonlinear susceptibility tensor ratio χ₃₃/χ₃₁ obtained from polarization‐resolved second harmonic generation (P‐SHG) under the assumption of cylindrical symmetry can be used to distinguish between fibrillar collagen types. Discriminating between collagen fibrils of types I and II is important in tissue engineering of cartilage. However, cartilage has a random organization of collagen fibrils, and the assumption of cylindrical symmetry may be incorrect. In this study, we simulated the P‐SHG response from different collagen organizations and demonstrated a possible method to exclude areas where cylindrical symmetry is not fulfilled and where fibrils are located in the imaging plane. The χ₃₃/χ₃₁‐ratio for collagen type I in tendon and collagen type II in cartilage was estimated to be 1.33 and 1.36, respectively, using this method. These ratios are now much closer than what has been reported previously in the literature, and the larger reported differences between collagen types can be explained by variation in the structural organization.      

Plasticity of proton pathway structure and water coordination in cytochrome c oxidase

Cytochrome c oxidase (CytcO) is a redox-driven, membrane-bound proton pump. One of the proton transfer pathways of the enzyme, the D pathway, used for the transfer of both substrate and pumped protons, accommodates a network of hydrogen-bonded water molecules that span the distance between an aspartate (Asp(132)), near the protein surface, and glutamate Glu(286), which is an internal proton donor to the catalytic site. To investigate how changes in the environment around Glu(286) affect the mechanism of proton transfer through the pathway, we introduced a non-hydrogen-bonding (Ala) or an acidic residue (Asp) at position Ser(197) (S197A or S197D), located approximately 7 A from Glu(286). Although Ser(197) is hydrogen-bonded to a water molecule that is part of the D pathway "proton wire," replacement of the Ser by an Ala did not affect the proton transfer rate. In contrast, the S197D mutant CytcO displayed a turnover activity of approximately 35% of that of the wild-type CytcO, and the O(2) reduction reaction was not linked to proton pumping. Instead, a fraction of the substrate protons was taken from the positive ("incorrect") side of the membrane. Furthermore, the pH dependence of the proton transfer rate was altered in the mutant CytcO. The results indicate that there is plasticity in the water coordination of the proton pathway, but alteration of the electrostatic potential within the pathway results in uncoupling of the proton translocation machinery.     

Seasonal changes in the hypothalamo-pituitary-gonadal axis in birds

Seasonal reproduction in birds is the adaptation to breed at the time of the greatest survival of young. This exact moment is mainly imposed by the photoperiod which stimulates the hypothalamo-pituitary-gonadal axis and starts breeding season. This article summarizes present knowledge concerning: 1/ perception and transduction of light into the biological signal; 2/ model of the avian photoperiodic response; 3/ seasonal changes in hypothalamic secretion of gonadotropin-releasing hormone (GnRH), vasoactive intestinal polipeptide (VIP), as well as the gonadotropin inhibitory hormone (GnIH); 4/ seasonal interactions between pituitary hormones; 5/ seasonal morphological and functional changes within the avian gonads.     

Specification of neuronal identities by feedforward combinatorial coding

Neuronal specification is often seen as a multistep process: earlier regulators confer broad neuronal identity and are followed by combinatorial codes specifying neuronal properties unique to specific subtypes. However, it is still unclear whether early regulators are re-deployed in subtype-specific combinatorial codes, and whether early patterning events act to restrict the developmental potential of postmitotic cells. Here, we use the differential peptidergic fate of two lineage-related peptidergic neurons in the Drosophila ventral nerve cord to show how, in a feedforward mechanism, earlier determinants become critical players in later combinatorial codes. Amongst the progeny of neuroblast 5–6 are two peptidergic neurons: one expresses FMRFamide and the other one expresses Nplp1 and the dopamine receptor DopR. We show the HLH gene collier functions at three different levels to progressively restrict neuronal identity in the 5–6 lineage. At the final step, collier is the critical combinatorial factor that differentiates two partially overlapping combinatorial codes that define FMRFamide versus Nplp1/DopR identity. Misexpression experiments reveal that both codes can activate neuropeptide gene expression in vast numbers of neurons. Despite their partially overlapping composition, we find that the codes are remarkably specific, with each code activating only the proper neuropeptide gene. These results indicate that a limited number of regulators may constitute a potent combinatorial code that dictates unique neuronal cell fate, and that such codes show a surprising disregard for many global instructive cues.     

Differential expression of duplicated LDH-A genes during temperature acclimation of weatherfish Misgurnus fossilis. Functional consequences for the enzyme

Temperature acclimation in poikilotherms entails metabolic rearrangements provided by variations in enzyme properties. However, in most cases the underlying molecular mechanisms that result in structural changes in the enzymes are obscure. This study reports that acclimation to low (5 degrees C) and high (18 degrees C) temperatures leads to differential expression of alternative forms of the LDH-A gene in white skeletal muscle of weatherfish, Misgurnus fossilis. Two isoforms of LDH-A mRNA were isolated and characterized: a short isoform (= 1332 bp) and a long isoform ( = 1550 bp), which both have 5'-UTRs and ORFs of the same length (333 amino acid residues), but differ in the length of the 3'-UTR. In addition, these two mRNAs have 44 nucleotide point mismatches of an irregular pattern along the complete sequence, resulting in three amino acid mismatches (Gly214Val; Val304Ile and Asp312Glu) between protein products from the short and long mRNA forms, correspondingly LDH-A(alpha) and LDH-A(beta) subunits. It is expected that the beta-subunit is more aliphatic due to the properties of the mismatched amino acids and therefore sterically more restricted. According to molecular modelling of M. fossilis LDH-A, the Val304Ile mismatch is located in the subunit contact area of the tetramer, whereas the remaining two mismatches surround the contact area; this is expected to manifest in the kinetic and thermodynamic properties of the assembled tetramer. In warm-acclimated fish the relative expression between alpha and beta isoforms of the LDH-A mRNA is around 5 : 1, whereas in cold-acclimated fish expression of is reduced almost to zero. This indicates that at low temperature the pool of total tetrameric LDH-A is more homogeneous in terms of alpha/beta-subunit composition. The temperature acclimation pattern of proportional pooling of subunits with different kinetic and thermodynamic properties of the tetrameric enzyme may result in fine-tuning of the properties of skeletal LDH-A, which is in line with previously observed kinetic and thermodynamic differences between 'cold' and 'warm' LDH-A purified from weatherfish. Also, an irregular pattern of nucleotide mismatches indicates that these mRNAs are the products of two independently evolving genes, i.e. paralogues. Karyotype analysis has confirmed that the experimental population of M. fossilis is tetraploid (2n = 100), therefore gene duplication, possibly through tetraploidy, may contribute to the adaptability towards temperature variation.     

The complex barnacle perfume: identification of waterborne pheromone homologues in Balanus improvisus and their differential expression during settlement

A key question in barnacle biology is the nature of cues that induce gregarious settlement. One of the characterised cues is the waterborne settlement pheromone (WSP). This study aimed to identify WSP homologues in Balanus improvisus and to investigate their expression during settlement. Six WSP homologues were identified, all containing an N-terminal signal peptide, a conserved core region, and a variable C-terminus comprising several -GR- and -HDDH- motifs. The B. improvisus WSP homologues were expressed in all settlement stages but showed different expression patterns. The homologue most similar to the B. amphitrite WSP was the most abundant and was constantly expressed during settlement. In contrast, several of the other WSP homologues showed the greatest expression in the juvenile stage. The presence of several WSP homologues suggests the existence of a pheromone mix, where con-specificity might be determined by a combination of sequence characteristics and the concentration of the individual components.     

Biochemical insights into basal and induced resistance in cabbage to black rot

Black rot, caused by Xanthomonas campestris pv. campestris (Xcc), is the most devastating disease of brassica, but the mechanisms of basal or induced resistance in cabbage remain largely unknown. Here, we performed three experiments to investigate biochemical features associated with cabbage resistance to black rot. In the first experiment, biochemical changes were assessed in plants that were inoculated with a highly (UFPR 5) or a moderately (Xcc 10) aggressive Xcc isolate. In the second experiment, we examined the biochemical responses in two cultivars (Chato de Quintal [CQ] and Louco de Verão [LV], susceptible and moderately resistant to Xcc, respectively). Finally, we examined whether acibenzolar‐S‐methyl (ASM) could induce cabbage resistance to Xcc. Plants inoculated with the Xcc 10 isolate displayed higher activities of superoxide dismutase (SOD), peroxidase (POX) and ascorbate peroxidase (APX), whereas activities of chitinase (CHI), β‐1,3‐glucanase (GLU) and polyphenol oxidase (PPO) as well as the concentrations of hydrogen peroxide (H₂O₂) and malondialdehyde (MDA) were lower compared to plants inoculated with the UFPR 5 isolate. The resistance of the cultivar LV to Xcc was linked to increases in the activities of CHI, GLU, and PPO and decreases in the activities of SOD, POX and APX as well as in the concentrations of H₂O₂ and MDA relative to the cultivar CQ. In general, ASM‐sprayed plants displayed higher activities for the enzymes studied, which was associated with decreased disease symptoms and oxidative stress. Taken together, our results demonstrated that high activities of both defence and antioxidant enzymes played a major role in both basal and induced resistance of cabbage to black rot.