Multiple actins in drosophila melanogaster

The tissue and developmental specificities of the three Drosophila isoactins, originally identified in primary myogenic cultures and in the permanent Schneider L-2 cell line, have been investigated. Of these three isoactins (I, II, and III), actins I and II are stable and actin III is unstable. Two-dimensional polyacrylamide gel electrophoretic analyses of total cellular extracts after 1-h [(35)S]methionine pulses were performed on a large variety of embryonic, larval, and adult muscle and nonmuscle tissues. The results suggest that isoactins II and III are generalized cellular actins found in all drosophila cell types. Actin I, on the other hand, is muscle-associated and is found exclusively in supercontractile muscle (such as larval body wall and larval and adult viscera) including primary myogenic cell cultures. Although actin I synthesis is not detectable during very early embryogenesis, it is detectable by 25 h and actin I is a major stable actin in all larval muscle tissues. Actin I is synthesized in reduced amounts relative to the other actins in late third instar larvae but is again a major product of actin synthesis in the adult abdomen. A stable actin species with the same pI as actin III has been identified in the adult thorax and appears to be unique to flight muscle tissue. This new stable form of thoracic actin may be the result of a stabilization of the actin III found in other tissues or may be an entirely separate gene product.     

Bloom's syndrome: DNA replication in cultured fibroblasts and lymphocytes

Summary Analysis of DNA fiber autoradiograms from Bloom's syndrome skin fibroblasts and blood lymphocytes shows a retarded rate of replication fork movement compared to normal adult controls. Other measurements from the autoradiograms—replication unit length, incidence of bidirectional replication, and degree of initiation synchrony—are normal in Bloom's syndrome cells. These results suggest that a slow rate of fork movement is a specific manifestation of defective DNA synthesis in all Bloom's syndrome cells.     

Functional characterization of PaLAX1, a putative auxin permease, in heterologous plant systems

We have isolated the cDNA of the gene PaLAX1 from a wild cherry tree (Prunus avium). The gene and its product are highly similar in sequences to both the cDNAs and the corresponding protein products of AUX/LAX-type genes, coding for putative auxin influx carriers. We have prepared and characterized transformed Nicotiana tabacum and Arabidopsis thaliana plants carrying the gene PaLAX1. We have proved that constitutive overexpression of PaLAX1 is accompanied by changes in the content and distribution of free indole-3-acetic acid, the major endogenous auxin. The increase in free indole-3-acetic acid content in transgenic plants resulted in various phenotype changes, typical for the auxin-overproducing plants. The uptake of synthetic auxin, 2,4-dichlorophenoxyacetic acid, was 3 times higher in transgenic lines compared to the wild-type lines and the treatment with the auxin uptake inhibitor 1-naphthoxyacetic acid reverted the changes caused by the expression of PaLAX1. Moreover, the agravitropic response could be restored by expression of PaLAX1 in the mutant aux1 plants, which are deficient in auxin influx carrier activity. Based on our data, we have concluded that the product of the gene PaLAX1 promotes the uptake of auxin into cells, and, as a putative auxin influx carrier, it affects the content and distribution of free endogenous auxin in transgenic plants.     

Identification of genetic factors influencing salt stress tolerance in white clover (Trifolium repens L.) by QTL analysis

Allotetraploid (2n = 4x = 32) white clover (Trifolium repens L.) is the most commonly cultivated legume component of temperate pastures, sown in swards with a companion grass species. Genetic control of growth performance of white clover on saline land is highly important for dairy industries, due to increasing soil salinity problems. The objective of this study was to identify quantitative trait loci (QTLs) for salinity tolerance in terms of vegetative growth under stress. Two parental genetic maps consisting of 213 and 159 marker loci and spanning 1,973.0 and 1,837.6 cM, respectively, were constructed using simple sequence repeat (SSR) and single nucleotide polymorphism (SNP) markers from a two-way pseudo-test cross F₁ population derived from pair-crossing of the Haifa₂ and LCL₂ genotypes. A total of 8 unique genomic regions on 8 linkage groups (LGs) of the Haifa₂ parental map and 6 unique regions on 5 LGs in the LCL₂ parental map were associated with plant growth under salt stress and relative growth under stress, as compared to control conditions. The results of this study indicate that salt tolerance in white clover is controlled by multiple QTLs, some at common locations, but each of limited magnitude. Location of these QTLs provides the genetic basis and potential for pyramiding of salt tolerance genes in breeding improvement.     

Combined Utilisation of Rapid Assessment Procedures for Loiasis (RAPLOA) and Onchocerciasis (REA) in Rain forest Villages of Cameroon

BACKGROUND: Individuals with high microfilarial loads of Loa loa are at increased risk of neurologic serious adverse (SAE) events following ivermectin treatment against onchocerciasis. RAPLOA (Rapid Assessment Procedure for loiasis), a newly developed rapid assessment procedure for loiasis that relates the prevalence of key clinical manifestation of loiasis (history of eye worm) to the level of endemicity of the infection (prevalence of high intensity), is a very useful tool to identify areas at potential risk of L. loa post ivermectin treatment encephalopathy. In a perspective of treatment decision making in areas of co-endemicity of loiasis/onchocerciasis, it would be advantageous (both in time and cost savings) for national onchocerciasis control programmes to use RAPLOA and the Rapid epidemiologic assessment for onchocerciasis (REA), in combination in given surveys. Since each of the two rapid assessment tools have their own specificities, the workability of combining the two methods needed to be tested. METHODS: We worked in 10 communities of a forest area presumed co-endemic for loiasis and onchocerciasis in the North-West Province of Cameroon where the mass-treatment with ivermectin had not been carried out. A four-step approach was used and comprised: (i) generating data on the prevalence and intensity of loiasis and onchocerciasis in an area where such information is scarce; (ii) testing the relationship between the L. loa microfilaraemia prevalence and the RAPLOA prevalence, (iii) testing the relationship between the O. volvulus microfiladermia prevalence and the REA prevalence, (iv) testing the workability of combining RAPLOA/REA by study teams in which a single individual can perform the interview for RAPLOA and the nodule palpation for REA. RESULTS: The microfilaraemia prevalence of loiasis in communities ranged from 3.6% to 14.3%. 6 (0.61%) individuals had L. loa microfilarial loads above 8000 mf/ml but none of them attained 30,000 mf/ml, the threshold value above which the risk of developing neurologic SAE after ivermectin treatment is very high. None of the communities surveyed had RAPLOA prevalence above 40%. All the communities had microfiladermia prevalence above 60%. The microfiladermia results could be confirmed by the rapid epidemiologic method (nodule palpation), with all the 10 communities having REA prevalence above 20%. For the first time, this study has demonstrated that the two rapid assessment procedures for loiasis and onchocerciasis can be carried out simultaneously by a survey team, in which a single individual can administer the questionnaire for RAPLOA and perform the nodule palpation for REA. CONCLUSION: This study has: (i) Revealed that the Momo valley of the North West province of Cameroon is hyperendemic for onchocerciasis, but is of lower level of endemicity for L. loa. (ii) Confirmed the previous relationships established between RAPLOA and the L. loa microfilaraemia prevalence in one hand and between the REA and the O. volvulus microfiladermia prevalence in another hand (iii) Shown that RAPLOA and REA could be used simultaneously for the evaluation of loiasis and onchocerciasis endemicity in areas targeted by the African Programme for onchocerciasis Control for community-directed treatment with ivermectin (CDTI).     

Regulation of meiotic prophase arrest in mouse oocytes by GPR3, a constitutive activator of the Gs G protein

The arrest of meiotic prophase in mouse oocytes within antral follicles requires the G protein G(s) and an orphan member of the G protein-coupled receptor family, GPR3. To determine whether GPR3 activates G(s), the localization of Galpha(s) in follicle-enclosed oocytes from Gpr3(+/+) and Gpr3(-/-) mice was compared by using immunofluorescence and Galpha(s)GFP. GPR3 decreased the ratio of Galpha(s) in the oocyte plasma membrane versus the cytoplasm and also decreased the amount of Galpha(s) in the oocyte. Both of these properties indicate that GPR3 activates G(s). The follicle cells around the oocyte are also necessary to keep the oocyte in prophase, suggesting that they might activate GPR3. However, GPR3-dependent G(s) activity was similar in follicle-enclosed and follicle-free oocytes. Thus, the maintenance of prophase arrest depends on the constitutive activity of GPR3 in the oocyte, and the follicle cell signal acts by a means other than increasing GPR3 activity.     

Nuclear envelope breakdown in starfish oocytes proceeds by partial NPC disassembly followed by a rapidly spreading fenestration of nuclear membranes

Breakdown of the nuclear envelope (NE) was analyzed in live starfish oocytes using a size series of fluorescently labeled dextrans, membrane dyes, and GFP-tagged proteins of the nuclear pore complex (NPC) and the nuclear lamina. Permeabilization of the nucleus occurred in two sequential phases. In phase I the NE became increasingly permeable for molecules up to approximately 40 nm in diameter, concurrent with a loss of peripheral nuclear pore components over a time course of 10 min. The NE remained intact on the ultrastructural level during this time. In phase II the NE was completely permeabilized within 35 s. This rapid permeabilization spread as a wave from one epicenter on the animal half across the nuclear surface and allowed free diffusion of particles up to approximately 100 nm in diameter into the nucleus. While the lamina and nuclear membranes appeared intact at the light microscopic level, a fenestration of the NE was clearly visible by electron microscopy in phase II. We conclude that NE breakdown in starfish oocytes is triggered by slow sequential disassembly of the NPCs followed by a rapidly spreading fenestration of the NE caused by the removal of nuclear pores from nuclear membranes still attached to the lamina.     

Classification for congenital anomalies of the hand: the IFSSH classification and the JSSH modification

The purpose of a classification for clinical problems which, except for a few specialized centers, occur only sporadically is to provide a system where these cases can be stored. This should allow all involved investigators to speak the same language; so-doing syndromes can be delinated, frequencies of occurence established and results of--different--treatments compared. A classification system should be simple to use, reliable and uniformly accepted. It should allow space for adaptations and/or extensions. The IFSSH proposed a 7 categories classification based on the proposed classification of Swanson et al. in 1976. This classification, was based on, which was thought in the seventies, etiopathogenic pathways. These 7 groups are: I. Failure of formation; transverse (A), or longitudinal (B) II. Failure of differentiation III. Polydactyly IV. Overgrowth V. Undergrowth VI. Amniotic band syndrome VII. Generalized skeletal syndromes. The extended classification proposed by IFSSH was used to classify 1013 hand differences in 925 hands of 650 patients. We found associated anomalies in 26.7%. The classification was straightforward in 86%, difficult in 6.6% and not possible in 7.8%. Group II was the most numerous group including 513 anomalies. We propose to include in this group the Madelung deformity, the Kirner deformity and congenital trigger fingers and trigger thumbs. In group I the radial and ulnar deficiencies, limited to the hand without forearm deficlencies should be Included. Triphalangeal thumbs are a problem, we suggest it to be listed in group III and consider it as a duplication in length. It is not always possible to evaluate the (transverse) absence of the fingers or hand. Longitudinal deficiencies (group IIB), symbrachydactyly (group V), and amniotic bands (group IV) occasionally develop a phenotype similar to the genuine transverse deficiency (group IA). Recently, the Japanese Society for Surgery of the Hand (JSSH) (16) proposed an extension/modification of the IFSSH classification. Based on newer knowledge on teratology, symbrachydactyly in all stages were transfered to group I. Two new groups were introduced. A group "failure of finger ray induction" including typical cleft hand (IC), central polydactyly (III) and (bony) syndactyly (II)--was included. Also a group of "unclassifiable" cases was added. This Japanese proposed classification is a real improvement and most clinicians and surgeons tend to use it in the future.     

Differences in isolated mitochondria are insufficient to account for respiratory depression during diapause in artemia franciscana embryos

In response to cues signifying the approach of winter, adult Artemia franciscana produce encysted embryos that enter diapause. We show that respiration rates of diapause embryos collected from the field (Great Salt Lake, Utah) are reduced up to 92% compared with postdiapause embryos when measured under conditions of normoxia and full hydration. However, mitochondria isolated from diapause embryos exhibit rates of state 3 and state 4 respiration on pyruvate that are equivalent to those from postdiapause embryos with active metabolism; a reduction in these rates (15%-27%) is measured with succinate for two of three collection years. Respiratory control ratios for diapause mitochondria are comparable to or higher than those from postdiapause embryos. The P : O flux ratios are statistically identical. Our calculations suggest that respiration of intact, postdiapause embryos is operating close to the state 3 oxygen fluxes measured for isolated mitochondria. Cytochrome c oxidase (COX) activity is 53% lower in diapause mitochondria during one collection year; the minimal impact of this COX reduction on mitochondrial respiration appears to be due to the 31% excess COX capacity in A. franciscana mitochondria. Transmission electron micrographs of embryos reveal mitochondria that are well differentiated and structurally similar in both states. As inferred from the similar amounts of mitochondrial protein extractable, tissue contents of mitochondria in diapause and postdiapause embryos are equivalent. Thus, metabolic depression during diapause cannot be fully explained by altered properties of isolated mitochondria. Rather, mechanisms for active inhibition or substrate limitation of mitochondrial metabolism in vivo may be operative.