Identification of Cell Cycle Dependent Interaction Partners of the Septins by Quantitative Mass Spectrometry

The septins are a conserved family of GTP-binding proteins that, in the baker''s yeast, assemble into a highly ordered array of filaments at the mother bud neck. These filaments undergo significant structural rearrangements during the cell cycle. We aimed at identifying key components that are involved in or regulate the transitions of the septins. By combining cell synchronization and quantitative affinity-purification mass-spectrometry, we performed a screen for specific interaction partners of the septins at three distinct stages of the cell cycle. A total of 83 interaction partners of the septins were assigned. Surprisingly, we detected DNA-interacting/nuclear proteins and proteins involved in ribosome biogenesis and protein synthesis predominantly present in alpha-factor arrested that do not display an assembled septin structure. Furthermore, two distinct sets of regulatory proteins that are specific for cells at S-phase with a stable septin collar or at mitosis with split septin rings were identified.Complementary methods like SPLIFF and immunoprecipitation allowed us to more exactly define the spatial and temporal characteristics of selected hits of the AP-MS screen.     

Oat mesophyll protoplasts: their response to various feeder cultures

Oat (Avena sativa L.) mesophyll protoplasts were recently demonstrated to be capable of dedifferentiation, repeated divisions, and colony formation. Since the development of oat mesophyll protoplasts is decisively influenced by the nature of the used feeder culture (species, variety and concentration), we conducted a systematic study of this parameter. Generally, graminaceous feeders promoted protoplast proliferation, while dicot species repressed protoplast divisions. The beneficial effect of those feeders that promote divisions was counterbalanced by a factor that causes necrosis. The correct balance between promotion of divisions or necrosis depended on the nature of the feeder and its plating density.     

Genetic and morphometric differentiation among island populations of two Norops lizards (Reptilia: Sauria: Polychrotidae) on independently colonized islands of the Islas de Bahia (Honduras)

Aim Anole lizards (Reptilia: Sauria: Polychrotidae) display remarkable morphological and genetic differentiation between island populations. Morphological differences between islands are probably due to both adaptive (e.g. differential resource exploitation and intra‐ or interspecific competition) and non‐adaptive differentiation in allopatry. Anoles are well known for their extreme diversity and rapid adaptive speciation on islands. The main aim of this study was to use tests of morphological and genetic differentiation to investigate the population structure and colonization history of islands of the Islas de Bahia, off the coast of Honduras. Location Five populations of Norops bicaorum and Norops lemurinus were sampled, four from islands of the Islas de Bahia and one from the mainland of Honduras. Methods Body size and weight differentiation were measured in order to test for significant differences between sexes and populations. In addition, individuals were genotyped using the amplified fragment length polymorphism technique. Bayesian model‐based and assignment/exclusion methods were used to study genetic differentiation between island and mainland populations and to test colonization hypotheses. Results Assignment tests suggested migration from the mainland to the Cayos Cochinos, and from there independently to both Utila and Roatán, whereas migration between Utila and Roatán was lacking. Migration from the mainland to Utila was inferred, but was much less frequent. Morphologically, individuals from Utila appeared to be significantly different in comparison with all other localities. Significant differentiation between males of Roatán and the mainland was found in body size, whereas no significant difference was detected between the mainland and the Cayos Cochinos. Main conclusions Significant genetic and morphological differentiation was found among populations. A stepping‐stone model for colonization, in combination with an independent migration to Utila and Roatán, was suggested by assignment tests and was compatible with the observed morphological differentiation.     

A note on the hatching and viability ofCeriodaphnia ephippia collected from lake sediment

Ephippia ofCeriodaphnia pulchella Sars were collected at 2 sites from successive sediment layers. Hatching observed in the laboratory gave information about the duration of their viability. Conclusions about the hatching situation in the lake were drawn from the ratio of intact to total ephippia at various lake depths. The results are discussed.     

Midbrain auditory sensitivity in the spring peeper (Pseudacris crucifer): correlations with behavioral studies

1. We derived audiograms from recordings of multiunit activity in the torus semicircularis of 10 males and 6 females of the spring peeper from central Missouri, USA. We used free-field stimulation with tone bursts that had temporal properties similar to typical advertisement calls and that ranged in frequency from 500-6000 Hz. 2. Audiograms from different electrode positions in the same animal had the same general shape. There was no evidence of tonotopy. 3. Audiograms showed two regions of maximal sensitivity: a low-frequency region (500-700 Hz); and a high-frequency region (2000-4000 Hz). Absolute thresholds and frequencies of maximum sensitivity varied considerably from individual to individual. 4. Audiograms derived from all individuals of each sex indicated that in the high-frequency region, corresponding to the frequency range of advertisement calls, males were more broadly tuned than females. However, tuning in both sexes was relatively weak, and the data predict relatively little selectivity in behavioral responses over the entire range of variation in frequency of the advertisement call in local populations. 5. The results are discussed in terms of behavioral experiments with both males and females from the same populations in central Missouri. We show that merely summarizing the audiograms based on estimates of minimum thresholds of a population or species may mask significant individual differences in tuning. Moreover, most behavioral studies are conducted at playback levels considerably above threshold. For these reasons, behavioral selectivity is not always accurately predicted by inspection of "average" audiograms.     

Assembly of 60S ribosomal subunits is perturbed in temperature-sensitive yeast mutants defective in ribosomal protein L16.

Temperature-sensitive mutants defective in 60S ribosomal subunit protein L16 of Saccharomyces cerevisiae were isolated through hydroxylamine mutagenesis of the RPL16B gene and plasmid shuffling. Two heat-sensitive and two cold-sensitive isolates were characterized. The growth of the four mutants is inhibited at their restrictive temperatures. However, many of the cells remain viable if returned to their permissive temperatures. All of the mutants are deficient in 60S ribosomal subunits and therefore accumulate translational preinitiation complexes. Three of the mutants exhibit a shortage of mature 25S rRNA, and one accumulates rRNA precursors. The accumulation of rRNA precursors suggests that ribosome assembly may be slowed in this mutant. These phenotypes lead us to propose that mutants containing the rpl16b alleles are defective for 60S subunit assembly rather than function. In the mutant carrying the rpl16b-1 allele, ribosomes initiate translation at the noncanonical codon AUA, at least on the rpl16b-1 mRNA, bringing to light a possible connection between the rate and the fidelity of translation initiation.     

Chemical characterization by protein sequence analysis of the bovine estrogen receptor

Tryptic peptides generated from bovine estrogen receptor have been fractionated and purified using microbore column high performance liquid chromatography. Sequence analysis performed on six of these peptides, derived from diverse structural regions of the receptor protein, yielded 73 unique assignments corresponding to approximately 12% of the molecule. The amino acid sequences of these peptides displayed a high degree of similarity with corresponding sequences from estrogen receptors of mammalian origin, but were only moderately conserved in receptors from non-mammalian species. The sequenced residues of one tryptic peptide, positioned in the estrogen binding domain, were fully conserved in all estrogen receptors.