Expression of lorelei-like genes in aposporous and sexual Paspalum notatum plants

Gametophytic apomictic plants form non-reduced embryo sacs that generate clonal embryos by parthenogenesis, in the absence of both meiosis and egg-cell fertilization. Here we report the sequence and expression analysis of a lorelei-like Paspalum notatum gene, n20gap-1, which encodes a GPI-anchored protein previously associated with apomixis in this species. Phylogeny trees showed that n20gap-1 was evolutionary related to the Arabidopsis thaliana lorelei genes At4g26466 and At5g56170. The lorelei At4g26466 disruption was shown to be detrimental to sperm cell release in arabidopsis. RFLP (Restriction Fragment Length Polymorphism) analysis revealed the occurrence of several homologous sequences in the Paspalum notatum genome, exhibiting polymorphisms genetically linked to apomixis. Real-time PCR showed that lorelei-family genes present a minor activity peak at pre-meiosis and a major one at anthesis. The apomictic genotype analyzed showed a significantly increased activity at pre-meiosis, post-meiosis and anthesis with respect to a sexual genotype. In situ hybridization assays revealed expression in integuments, nucellus and the egg-cell apparatus. Several n20gap-1 alleles differing mainly at the 3' UTR sequence were identified. Allele-specific real-time PCR experiments showed that allele 28 was significantly induced in reproductive tissues of the apomictic genotype with respect to the sexual genotype at anthesis. Our results indicate that P. notatum lorelei-like genes are differentially expressed in representative sexual (Q4188) and apomictic (Q4117) genotypes, and might play a role in the final stages of the apomixis developmental cascade. However, the association of n20gap-1 expression with the trait should be confirmed in significant number of sexual and apomictic genotypes.     

Ohio USA stoneflies (Insecta, Plecoptera): species richness estimation, distribution of functional niche traits, drainage affiliations, and relationships to other states

Ohio is an eastern USA state that historically was >70% covered in upland and mixed coniferous forest; about 60% of it glaciated by the Wisconsinan glacial episode. Its stonefly fauna has been studied in piecemeal fashion until now. The assemblage of Ohio stoneflies was assessed from over 4,000 records accumulated from 18 institutions, new collections, and trusted literature sources. Species richness totaled 102 with estimators Chao2 and ICE Mean predicting 105.6 and 106.4, respectively. Singletons and doubletons totaled 18 species. All North American families were represented with Perlidae accounted for the highest number of species at 34. The family Peltoperlidae contributed a single species. Most species had univoltine-fast life cycles with the vast majority emerging in summer, although there was a significant component of winter stoneflies. Nine United States Geological Survey hierarchical drainage units level 6 (HUC6) were used to stratify specimen data. Species richness was significantly related to the number of unique HUC6 locations, but there was no relationship with HUC6 drainage area. A nonparametric multidimensional scaling analysis found that larger HUC6s in the western part of the state had similar assemblages with lower species richness that were found to align with more savanna and wetland habitat. Other drainages having richer assemblages were aligned with upland deciduous and mixed coniferous forests of the east and south where slopes were higher. The Ohio assemblage was most similar to the well-studied fauna of Indiana (88 spp.) and Kentucky (108 spp.), two neighboring states. Many rare species and several high quality stream reaches should be considered for greater protection.     

Contributions of the hippocampus and medial prefrontal cortex to energy and body weight regulation

The effects of selective ibotenate lesions of the complete hippocampus (CHip), the hippocampal ventral pole (VP), or the medial prefrontal cortex (mPFC) in male rats were assessed on several measures related to energy regulation (i.e., body weight gain, food intake, body adiposity, metabolic activity, general behavioral activity, conditioned appetitive responding). The testing conditions were designed to minimize the nonspecific debilitating effects of these surgeries on intake and body weight. Rats with CHip and VP lesions exhibited significantly greater weight gain and food intake compared with controls. Furthermore, CHip-lesioned rats, but not rats with VP lesions, showed elevated metabolic activity, general activity in the dark phase of the light-dark cycle, and greater conditioned appetitive behavior, compared with control rats without these brain lesions. In contrast, rats with mPFC lesions were not different from controls on any of these measures. These results indicate that hippocampal damage interferes with energy and body weight regulation, perhaps by disrupting higher-order learning and memory processes that contribute to the control of appetitive and consummatory behavior.     

Identification of a maize linkage group related to apomixis in Brachiaria

 A bulked segregant analysis using RFLPs and RAPDs was carried out to identify molecular markers co-segregating with apomixis in a Brachiaria F₁ population. The test population used was a cross between sexual B. ruziziensis R44 and the aposporous apomictic Brachiaria brizantha cv Marandu. The Brachiaria genome was systematically scanned using 61 cDNA and genomic maize clones detecting 65 loci located at 40 cM, on average, one from each other in the maize genome. The finding of a clone that presented a polymorphic band co-segregating with apomixis (umc147) led to the identification of another marker within the same area (umc72). The clones belong to a duplicated linkage group that maps to the distal part of maize chromosome-1 long arm and chromosome-5 short arm. RAPD analysis using 184 primers from Operon sets yielded one more marker (OPC4) significantly linked to the trait mapping the same locus. OPC4 had been previously reported as a potential marker for apospory in Pennisetum. A map of the region was constructed using additional clones that belong to the same maize linkage group. Since that was the only genomic region that presented an apomixis-linked polymorphism our observations support the existence of a single locus directing apospory in Brachiaria. Received: 9 September 1996 / Accepted: 20 September 1996     

Evidence that functional erythrocyte-type glucose transporters are oligomers

In this study we tested the hypothesis that functional erythrocyte-type glucose transporters (GLUT1) exist as oligomeric complexes by expressing chimeric transporter proteins in Chinese hamster ovary cells harboring endogenous GLUT1 transporters. The chimeric transporters were GLUT1-4c, in which the 29 C-terminal residues of human GLUT1 were replaced by the 30 C-terminal residues of rat skeletal muscle glucose transporter (GLUT4), and GLUT1n-4, containing the N-terminal 199 residues of GLUT1 and the 294 C-terminal residues of GLUT4. Endogenous GLUT1 was quantitatively co-immunoprecipitated by using an anti-GLUT4 C-terminal peptide antibody from detergent extracts of Chinese hamster ovary cells expressing either of the chimeric proteins, as detected by immunoblotting the precipitates with an anti-GLUT1 C-terminal peptide antiserum. No co-immunoprecipitation of native GLUT1 with native GLUT4 from extracts of 3T3-L1 adipocytes, which contain both these transporters, was observed with the same antibody. These data are consistent with the hypothesis that GLUT1 transporters exist as homodimers or higher order oligomers and that a major determinant of oligomerization is located within the first 199 residues of GLUT1.     

Positive selection driving the evolution of a gene of male reproduction, Acp26Aa, of Drosophila: II. Divergence versus polymorphism

The evolution of the gene for a male ejaculatory protein, Acp26Aa, has been shown to be driven by positive selection when nonsibling species in the Drosophila melanogaster subgroup are compared. To know if selection has been operating in the recent past and to understand the details of its dynamics, we obtained DNA sequences of Acp26Aa and the nearby Acp26Ab gene from 39 D. melanogaster chromosomes. Together with the 10 published sequences, we analyzed 49 sequences from five populations in four continents. The southern African population is somewhat differentiated from all other populations, but its nucleotide diversity is lower at these two loci. We find the following results for Acp26Aa: (1) The R: S (replacement : silent changes) ratio is significantly higher in the between-species comparisons than in the within-species data by the McDonald and Kreitman test. Positive selection is probably responsible for the excess of amino acid replacements between species. (2) However, within-species nucleotide diversity is high. Neither the Tajima test nor the Fu and Li test indicates a reduction in nucleotide diversity due to positive selection in the recent past. (3) The newly derived nucleotides in D. melanogaster are at high frequency significantly more often than predicted by the neutral equilibrium. Since the nearby Acp26Ab gene does not show these patterns, these observations cannot be attributed to the characteristics of this chromosomal region. We suggest that positive selection is active, but may be weak, for each amino acid change in the Acp26Aa gene.