A chromosome-level genome assembly enables the identification of the follicule stimulating hormone receptor as the master sex-determining gene in the flatfish Solea senegalensis

Sex determination (SD) shows huge variation among fish and a high evolutionary rate, as illustrated by the Pleuronectiformes (flatfishes). This order is characterized by its adaptation to demersal life, compact genomes and diversity of SD mechanisms. Here, we assembled the Solea senegalensis genome, a flatfish of great commercial value, into 82 contigs (614 Mb) combining long- and short-read sequencing, which were next scaffolded using a highly dense genetic map (28,838 markers, 21 linkage groups), representing 98.9% of the assembly. Further, we established the correspondence between the assembly and the 21 chromosomes by using BAC-FISH. Whole genome resequencing of six males and six females enabled the identification of 41 single nucleotide polymorphism variants in the follicle stimulating hormone receptor (fshr) consistent with an XX/XY SD system. The observed sex association was validated in a broader independent sample, providing a novel molecular sexing tool. The fshr gene displayed differential expression between male and female gonads from 86 days post-fertilization, when the gonad is still an undifferentiated primordium, concomitant with the activation of amh and cyp19a1a, testis and ovary marker genes, respectively, in males and females. The Y-linked fshr allele, which included 24 nonsynonymous variants and showed a highly divergent 3D protein structure, was overexpressed in males compared to the X-linked allele at all stages of gonadal differentiation. We hypothesize a mechanism hampering the action of the follicle stimulating hormone driving the undifferentiated gonad toward testis.     

How did glycogen structure evolve to satisfy the requirement for rapid mobilization of glucose? A problem of physical constraints in structure building

Optimization of molecular design in cellular metabolism is a necessary condition for guaranteeing a good structure–function relationship. We have studied this feature in the design of glycogen by means of the mathematical model previously presented that describes glycogen structure and its optimization function [Meléndez-Hevia et al. (1993), Biochem J 295: 477–483]. Our results demonstrate that the structure of cellular glycogen is in good agreement with these principles. Because the stored glucose in glycogen must be ready to be used at any phase of its synthesis or degradation, the full optimization of glycogen structure must also imply the optimization of every intermediate stage in its formation. This case can be viewed as a molecular instance of the eye problem, a classical paradigm of natural selection which states that every step in the evolutionary formation of a functional structure must be functional. The glycogen molecule has a highly optimized structure for its metabolic function, but the optimization of the full molecule has meaning and can be understood only by taking into account the optimization of each intermediate stage in its formation. Received: 23 October 1996 / Accepted: 21 April 1997     

Acutely administered melatonin reduces oxidative damage in lung and brain induced by hyperbaric oxygen

Pablos, Marta I., Russel J. Reiter, Jin-Ing Chuang, GenaroG. Ortiz, Juan M. Guerrero, Ewa Sewerynek, Maria T. Agapito, DanielaMelchiorri, Richard Lawrence, and Susan M. Deneke. Acutely administered melatonin reduces oxidative damage in lung and brain induced by hyperbaric oxygen. J. Appl.Physiol. 83(2): 354-358, 1997.Hyperbaric oxygenexposure rapidly induces lipid peroxidation and cellular damage in avariety of organs. In this study, we demonstrate that the exposure ofrats to 4 atmospheres of 100% oxygen for 90 min is associated withincreased levels of lipid peroxidation products [malonaldehyde(MDA) and 4-hydroxyalkenals (4-HDA)] and withchanges in the activities of two antioxidative enzymes[glutathione peroxidase (GPX) and glutathione reductase (GR)], as well as in the glutathione status in the lungs and in the brain. Products of lipid peroxidation increased after hyperbaric hyperoxia, both GPX and GR activities were decreased, and levels oftotal glutathione (reduced+oxidized) and glutathione disulfide (oxidized glutathione) increased in both lung and brain areas (cerebralcortex, hippocampus, hypothalamus, striatum, and cerebellum) but not inliver. When animals were injected with melatonin (10 mg/kg) immediatelybefore the 90-min hyperbaric oxygen exposure, all measurements ofoxidative damage were prevented and were similar to those in untreatedcontrol animals. Melatonin's actions may be related to a variety ofmechanisms, some of which remain to be identified, including itsability to directly scavenge free radicals and its induction ofantioxidative enzymes via specific melatonin receptors.

     

Sequences isolated from aDrosophila early-ecdysone puff are expressed in rat liver

We have studied the presence of a cloned fragment of DNA from Drosophila melanogaster in other organisms by means of nucleic acid hybridization analysis. The isolated region is localized in polytene chromosomes at the 63F subdivision. This region includes a puff that responds within minutes to ecdysone stimulation. We have found that 63F DNA from D. melanogaster hybridizes 'in situ' to both DNA and RNA from D. simulans, D. teissieri, and D. hydei. In all these species the isolated DNA remains associated with one early-ecdysone stimulated puff. The isolated Drosophila recombinant DNA is also complementary to polyadenylated RNA from foetal and adult rat liver but fails to hybridize to the nonpolyadenylated RNA classes from both sources and to polyadenylated RNA from rat mammary glands.     

Loose binding of testicular mitochondrial ATPase to the inner membrane

Rat testis mitochondrial ATPase was not inhibited by oligomycin at pH 7.5. It was inhibited only at higher alkaline pH's, and showed a lower sensitivity both to oligomycin and N,N′-dicyclohexylcarbodiimide and a higher one to efrapeptin. In submitochondrial particles, testis ATPase was only slightly inhibited by oligomycin, ossamycin, and efrapeptin. The possibility of a loose binding of F₁ to the membrane was supported by its recovery from the supernatant of the submitochondrial particles. Furthermore, by electron microscopy, after hypoosmotic shock and negative staining of the mitochondrial preparations, most of the inner mitochondrial membranes showed only a few “knobs” or none at all. The capacity of the testis mitochondrial preparation to produce ATP was tested and compared to that from liver. ATP synthetase/ATPase activity ratio was $\text{30}\text{1}$ in liver mitochondria, whereas in the testis it was $\text{3}\text{1}$. In spite of this large difference, at least part of the testis ATPase must be firmly bound to the membrane, since it is able to form ATP. The rest seems to be loosely bound and its functional significance is still unknown.     

Effect of rye embryo ribosome nuclease on double-stranded RNA

In extracts obtained by treating rye embryo ribosomes with 0.5 M NH₄Cl, nuclease activity was noted towards double-stranded RNA from virus of Penicillium chrysogenum and towards synthetic poly (A)-poly (U) and poly (I)-poly (C) complexes.     

Cell free synthesis of some storage protein subunits by polyribosomes and RNA isolated from developing seeds of pea (Pisum sativum L.)

Polyribosomes which have template activity in the wheat germ system have been isolated from developing pea seeds. Some of the translation products have identical mobilities to the vicilin and legumin subunits by SDS-PAGE. Certain products were specifically immunoprecipitated with antisera prepared against purified vicilin and legumin fractions. Various RNA fractions including poly A-rich RNA have also been isolated from polyribosomes and shown to direct the synthesis of polyripeptides whose properties are similar to the storage protein subunits. The results are discussed in relationship to other investigations with seed storage protein biosynthesis in vitro.Abbreviations DTT dithiothreitol - SDS-PAGE SDS-polyacrylamide gel electrophoresis - TCA tricarboxylic acid     

CAPABILITY OF TUBULIN AND MICROTUBULES TO INCORPORATE AND TO RELEASE TYROSINE AND PHENYLALANINE AND THE EFFECT OF THE INCORPORATION OF THESE AMINO ACIDS ON TUBULIN ASSEMBLY

Abstract— Incorporation of [¹⁴C]tyrosine into the C-terminal position of α-tubulin of rat brain cytosol was 10-fold higher for non-assembled than for assembled tubulin. The incorporation into tubulin from disassembled microtubules was higher than into non-assembled tubulin; therefore, the low incorporation into microtubules was not due to a lower acceptor capacity of their tubulin constituent.
[¹⁴C]Tyrosine was released from assembled and non-assembled [¹⁴C]tyrosinated tubulin by the action of an endogenous carboxypeptidase. Release from non-assembled tubulin was shown by incubating a tubulinyl-[¹⁴C]tyrosine preparation in the presence of CaCl₂ at a concentration that abolished microtubule formation. Release from microtubules was inferred from the observation that the percentages of [¹⁴C]tyrosine released and the decrease of the specific radioactivity of the recovered microtubules were practically identical and did not change after a 10-fold dilution of the incubated microtubules.
[³H]Phenylalanine was released from a preparation of tubulinyl-[³H]phenylalanine also by an enzymatic activity.
The capacity of a tubulin preparation to incorporate tyrosine was increased 43% by pre-treatment with endogenous carboxypeptidase.
Tubulin tyrosinated in vitro was assembled to the same extent as native tubulin. After a mixture of tubulinyl-[¹⁴C]tyrosine and tubulinyl-[³H]phenylalanine was partially assembled, the ratio of ¹⁴C/³H found in the microtubules was the same as in the non-assembled tubulin fraction.