RESUMEN
Dysferlin is a Ca(2+)-binding protein found in many different cell types. It is required for membrane wound repair in muscle, but it is not known whether it has the same function in other cells. Here we report the activation of an intercellular signaling pathway in sea urchin embryos by membrane wounding that evokes Ca(2+) spikes in neighboring cells. This pathway was mimicked by ATP application, and inhibited by apyrase, cadmium, and omega-agatoxin-IVA. Microinjection of dysferlin antisense phosphorodiamidate morpholino oligonucleotides blocked this pathway, whereas control morpholinos did not. Co-injection of mRNA encoding human dysferlin with the inhibitory morpholino rescued signaling activity. We conclude that in sea urchin embryos dysferlin mediates Ca(2+)-triggered intercellular signaling in response to membrane wounding.
Asunto(s)
Adenosina Trifosfato/metabolismo , Membrana Celular/metabolismo , Proteínas de la Membrana/metabolismo , Proteínas Musculares/metabolismo , Animales , Apirasa/farmacología , Cadmio/farmacología , Señalización del Calcio/efectos de los fármacos , Señalización del Calcio/genética , Membrana Celular/efectos de los fármacos , Disferlina , Células HeLa , Humanos , Inmunohistoquímica , Rayos Láser , Proteínas de la Membrana/genética , Microscopía de Fluorescencia por Excitación Multifotónica , Proteínas Musculares/genética , Oligonucleótidos Antisentido/genética , Erizos de Mar , Cicatrización de Heridas/genéticaRESUMEN
Eukaryotic cells have multiple forms of endocytosis which maintain cell surface homeostasis. One explanation for this apparent redundancy is to allow independent retrieval of surface membranes derived from different types of vesicles. Consistent with this hypothesis we find that sea urchin eggs have at least two types of compensatory endocytosis. One is associated with retrieving cortical vesicle membranes, and formed large endosomes by a mechanism that was inhibited by agatoxin, cadmium, staurosporine and FK506. The second type is thought to compensate for constitutive exocytosis, and formed small endosomes using a mechanism that was insensitive to the above mentioned reagents, but was inhibited by phenylarsine oxide (PAO), and by microinjection of mRNA encoding Src kinase. Both mechanisms could act concurrently, and account for all of the endocytosis occurring during early development. Inhibition of either form did not trigger compensation by the other form, and phorbol ester treatment rescued the endocytotic activity blocked by agatoxin, but not the retrieval blocked by PAO.
