Domenico Cascino

A constitutive 70 kDa heat-shock protein is localized on the fibres of spindles and asters at metaphase in an ATP-dependent manner: a new chaperone role is proposed.

In the present study, double immunofluorescence and immunoblot analysis have been used to show that centrosomes, isolated from Paracentrotus lividus sea urchin embryos at the first mitotic metaphase, contain the constitutive chaperone, heat-shock protein (HSP) 70. More specifically, we demonstrate that centrosomes contain only the HSP70-d isoform, which is one of the four isoforms identified in P. lividus. We also provide evidence that p34(cell division control kinase-2) and t complex polypeptide-1 (TCP-1) alpha, a subunit of the TCP-1 complex, are localized on the centrosomes. Furthermore, inhibition of TCP-1 in vivo, via microinjecting an anti-(TCP-1 alpha) antibody into P. lividus eggs before fertilization, either impaired mitosis or induced severe malformations in more than 50% of embryos. In addition, we have isolated the whole mitotic apparatus and shown that HSP70 localizes on the fibres of spindles and asters, and binds them in an ATP-dependent manner. These observations suggest that HSP70 has a chaperone role in assisting the TCP-1 complex in tubulin folding, when localized on centrosomes, and during the assembling and disassembling of the mitotic apparatus, when localized on the fibres of spindles and asters.

Myo-inositol counteracts the vegetalizing effect of lithium on P.lividus embryos

Abstract The vegetalizing effect of LiCl on sea urchins embryos can be counteracted by the addition of myo-inositol. This observation is discussed in connection with similar results recently reported for amphibian embryos.

EFFECT OF THE IMPase INHIBITOR L690,330 ON SEA URCHIN DEVELOPMENT

A variety of concentrations of the IMPase inhibitor L690,330 were added to sea urchin embryos. Immediate arrest of development was obtained for concentrations from 7.5mmon. Concentrations lower than 3.5mmpermitted gastrulation but inhibited skeletogenesis and disturbed elongation along the animal—vegetal axis. The latter results are similar to those obtained by counteracting lithium effect with myoinositol, which are suggested to be due to partial relief of IMPase inhibition.

Achievement of thermotolerance trough hsps phosphorylation in sea urchin embryos.

TPA treatment of sea urchin embryos is able to induce thermotolerance. Evidence is provided that TPA treatment induces phosphorylation of a constitutive stress protein of 38 KDa.

Determination of the sequence and study of the expression of a gene for the heat shock protein (hsp) 70 of the sea urchin Paracentrotus lividus

The sequence of a gene coding for the hsp70 isolated from a P.lividus genomic library is reported. This gene is interrupted by a long intron at 183 nt downstream the translation start codon. Its promoter region contains three heat shock elements, thus indicating that the gene is an inducible one. Northern blot analysis shows that this gene is transcribed following heat shock and that its transcription products bind to ribosomes.RiassuntoSi riporta la sequenza nucleotidica di un gene per la proteina da shock da calore di 70 kDa isolato da una genoteca di riccio di mareParacentrotus lividus. Quesro gene è interrotto da un lungo introne a 183 nucleotidi a valle della tripletta per l’inizio della traduzione. La sua regione del promotore contiene tre elementiheat shock, il ehe indica ehe trattasi di un gene inducibile. Un’analisi perNorthern blot mostra ehe il gene è trascritto a seguito di shock da calore e che i prodotti della sua trascrizione si legano ai ribosomi.