Evolution of the 28S ribosomal RNA gene in anurans: regions of variability and their phylogenetic implications

Fifteen restriction sites were mapped to the 28S ribosomal RNA gene of individuals representing 54 species of frogs, two species of salamanders, a caecilian, and a lungfish. Eight of these sites were present in all species examined, and two were found in all but one species. Alignment of these conserved restriction sites revealed, among anuran 28S rRNA genes, five regions of major length variation that correspond to four of 12 previously identified divergent domains of this gene. One of the divergent domains (DD8) consists of two regions of length variation separated by a short segment that is conserved at least throughout tetrapods. Most of the insertions, deletions, and restriction-site variations identified in the 28S gene will require sequence-level analysis for a detailed reconstruction of their history. However, an insertion in DD9 that is coextensive with frogs in the suborder Neobatrachia, a BstEII site that is limited to representatives of two leptodactylid subfamilies, and a deletion in DD10 that is found only in three ranoid genera are probably synapomorphies.      

Characterization of cross-bridge elasticity and kinetics of cross-bridge cycling during force development in single smooth muscle cells

Force development in smooth muscle, as in skeletal muscle, is believed to reflect recruitment of force-generating myosin cross-bridges. However, little is known about the events underlying cross-bridge recruitment as the muscle cell approaches peak isometric force and then enters a period of tension maintenance. In the present studies on single smooth muscle cells isolated from the toad (Bufo marinus) stomach muscularis, active muscle stiffness, calculated from the force response to small sinusoidal length changes (0.5% cell length, 250 Hz), was utilized to estimate the relative number of attached cross-bridges. By comparing stiffness during initial force development to stiffness during force redevelopment immediately after a quick release imposed at peak force, we propose that the instantaneous active stiffness of the cell reflects both a linearly elastic cross-bridge element having 1.5 times the compliance of the cross-bridge in frog skeletal muscle and a series elastic component having an exponential length-force relationship. At the onset of force development, the ratio of stiffness to force was 2.5 times greater than at peak isometric force. These data suggest that, upon activation, cross-bridges attach in at least two states (i.e., low-force-producing and high-force-producing) and redistribute to a steady state distribution at peak isometric force. The possibility that the cross-bridge cycling rate was modulated with time was also investigated by analyzing the time course of tension recovery to small, rapid step length changes (0.5% cell length in 2.5 ms) imposed during initial force development, at peak force, and after 15 s of tension maintenance. The rate of tension recovery slowed continuously throughout force development following activation and slowed further as force was maintained. Our results suggest that the kinetics of force production in smooth muscle may involve a redistribution of cross-bridge populations between two attached states and that the average cycling rate of these cross-bridges becomes slower with time during contraction.     

Evidence for "twisted plane" undulations in golden hamster sperm tails

Motile spermatozoa from the golden hamster have been arrested by rapid freezing and then fixed with glutaraldehyde at low temperature after substitution with ethylene glycol. As far as can be judged, the flagellar waveforms thus stabilized are similar to those seen in living sperm; in contrast, fixation in glutaraldehyde, without prior freezing, induces agonal changes in flagellar conformation. The characteristics waveform after freeze substitution contains three bends. Approx. half of these flagella are entirely planar. The rest are three dimensional, with the third bend displaced in a regular way from the plane containing the proximal two bends. From the geometry of these flagella, it is concluded that the plane of action of a given bending cycle undergoes a clockwise twist (from a forward viewpoint) as the cycle is succeeded by new bending cycles. This "twisted plane" undulation is quite different from helical movement. The twisting seems to occur abruptly, between cycles, as if each bending cycle has a preferred plane of action. The mechanism underlying the twisting is uncertain. However, on the basis of the angular displacements between the preferred planes, and the findings from electron microscopy, the following idea is presented as a working hypothesis: that, if the most proximal plane of bending is topographically determined by peripheral doublet 1, then successive distal planes of action are influenced predominantly by doublets 2, 3, etc., in clockwise sequence. The merits and weaknesses of this hypothesis are discussed.     

Variation in the ribosomal internal transcribed spacers and 5.8S rDNA among five species of Acropora (Cnidaria; Scleractinia): patterns of variation consistent with reticulate evolution

The ITS sequences of Acropora spp. are the shortest so far identified in any metazoan and are among the shortest seen in eukaryotes; ITS1 was 70-80 bases, and ITS2 was 100-112 bases. The ITS sequences were also highly variable, but base composition and secondary structure prediction indicate that divergent sequence variants are unlikely to be pseudogenes. The pattern of variation was unusual in several other respects: (1) two distinct ITS2 types were detected in both A. hyacinthus and A. cytherea, species known to hybridize in vitro with high success rates, and a putative intermediate ITS2 form was also detected in A. cytherea; (2) A. valida was found to contain highly (29%) diverged ITS1 variants; and (3) A. longicyathus contained two distinct 5.8S rDNA types. These data are consistent with a reticulate evolutionary history for the genus Acropora.      

Teaching Bernoulli's Principle through Demonstrations

ABSTRACT

One proven strategy to help students make sense of abstract concepts is to sequence instruction so students have exploratory opportunities to investigate science before being introduced to new science explanations (Abraham and Renner 1986 Abraham, M. R. and Renner, J. W. 1986. The sequence of learning cycle activities in high school chemistry. Journal of Research in Science Teaching, 23: 121–143. [Crossref], [Web of Science ®] , [Google Scholar]; Renner, Abraham, and Birnie 1988 Renner, J. W., Abraham, M. R. and Birnie, H. H. 1988. The necessity of each phase of the learning cycle in teaching high school physics. Journal of Research in Science Teaching, 25: 39–58. [Crossref], [Web of Science ®] , [Google Scholar]). To help physical science teachers make sense of how to effectively sequence lessons, this article summarizes our experiences using an exploration–explanation sequence of instruction to teach Bernoulli's principle to prospective middle and secondary science teachers in a science methods course. We use demonstrations during our Bernoulli unit to help students go back and forth between their observations of phenomenon and what occurs on the microscopic level with what we have termed molecular talk. Students engage in guiding questions, consider their old and new understandings of science, and use evidence to construct new ideas during all stages of the lesson.     

Isocitrate dehydrogenase 1 R132C mutation occurs exclusively in microsatellite stable colorectal cancers with the CpG island methylator phenotype

The CpG Island Methylator Phenotype (CIMP) is fundamental to an important subset of colorectal cancer; however, its cause is unknown. CIMP is associated with microsatellite instability but is also found in BRAF mutant microsatellite stable cancers that are associated with poor prognosis. The isocitrate dehydrogenase 1 (IDH1) gene causes CIMP in glioma due to an activating mutation that produces the 2-hydroxyglutarate oncometabolite. We therefore examined IDH1 alteration as a potential cause of CIMP in colorectal cancer. The IDH1 mutational hotspot was screened in 86 CIMP-positive and 80 CIMP-negative cancers. The entire coding sequence was examined in 81 CIMP-positive colorectal cancers. Forty-seven cancers varying by CIMP-status and IDH1 mutation status were examined using Illumina 450K DNA methylation microarrays. The R132C IDH1 mutation was detected in 4/166 cancers. All IDH1 mutations were in CIMP cancers that were BRAF mutant and microsatellite stable (4/45, 8.9%). Unsupervised hierarchical cluster analysis identified an IDH1 mutation-like methylation signature in approximately half of the CIMP-positive cancers. IDH1 mutation appears to cause CIMP in a small proportion of BRAF mutant, microsatellite stable colorectal cancers. This study provides a precedent that a single gene mutation may cause CIMP in colorectal cancer, and that this will be associated with a specific epigenetic signature and clinicopathological features.     

Carboxylesterase 1 Gene Duplication and mRNA Expression in Adipose Tissue Are Linked to Obesity and Metabolic Function

Context and Aims

Carboxylesterase 1 (CES1) appears to play an important role in the control of the metabolism of triglycerides and cholesterol in adipocytes and other cell types including hepatocytes. Therefore, it is relevant to gain insights into the genetic versus non-genetic mechanisms involved in the control of CES1 mRNA expression. Here, we investigated CES1 mRNA expression level in adipose tissue and its association with measures of adiposity and metabolic function in a population of elderly twins. Furthermore, the heritability of CES1 mRNA expression level in adipose tissue and the effect of CES1 gene duplication were assessed.

Methodology

A total of 295 monozygotic and dizygotic twin subjects (62–83 years) with (n = 48) or without (n = 247) type 2 diabetes mellitus were enrolled in the study. They were subjected to a standard oral glucose tolerance test and excision of abdominal subcutaneous fat biopsies during the fasting state. Levels of CES1 mRNA and copy number of the gene were assessed by quantitative PCR.

Results

CES1 mRNA expression level in adipose tissue was positively associated with body-mass index (P<0.001), homeostasis model assessment-insulin resistance (P = 0.003) and level of fasting glucose (P = 0.002), insulin (P = 0.006), and triglycerides (P = 0.003). The heritability for the expression of CES1 mRNA in adipose tissue was high. CES1 gene duplication was positively associated with insulin sensitivity (P = 0.05) as well as glucose tolerance (P = 0.03) and negatively associated with homeostasis model assessment-insulin resistance (P = 0.02). Duplication of CES1 was not linked to mRNA level of this gene (P = 0.63).

Conclusion

CES1 mRNA in adipose tissue appears to be under strong genetic control and was associated with measures of metabolic function raising the possibility of a potential role of this enzyme in the development of type 2 diabetes mellitus. Further studies are needed to understand the potential effect of CES1 gene duplication on adipocyte and whole-body metabolic functions.     

The long-acting glucagon-like peptide-1 analogue, liraglutide, inhibits beta-cell apoptosis in vitro

We here show that GLP-1 and the long-acting GLP-1 analogue, liraglutide, interfere with diabetes-associated apoptotic processes in the β-cell. Studies using primary neonatal rat islets showed that native GLP-1 and liraglutide inhibited both cytokine- and free fatty acid-induced apoptosis in a dose-dependent manner. The anti-apoptotic effect of liraglutide was mediated by the GLP-1 receptor as the specific GLP-1 receptor antagonist, exendin(9-39), blocked the effects. The adenylate cyclase activator, forskolin, had an anti-apoptotic effect similar to those of GLP-1 and liraglutide indicating that the effect was cAMP-mediated. Blocking the PI3 kinase pathway using wortmannin but not the MAP kinase pathways by PD98059 inhibited the effects of liraglutide. In conclusion, GLP-1 receptor activation has anti-apoptotic effect on both cytokine, and free fatty acid-induced apoptosis in primary islet-cells, thus suggesting that the long-acting GLP-1 analogue, liraglutide, may be useful for retaining β-cell mass in both type 1 and type 2 diabetic patients.