An Ultrastructural Investigation on Vitellophage Invasion of the Yolk Mass during and after Germ Band Formation in Embryos of the Stick Insect <i>Carausius morosus Br.</i>

Fausto, Anna Maria; Carcupino, Marcella; Mazzini, Massimo; Giorgi, Franco

doi:10.1111/j.1440-169x.1994.00197.x

Cited by 16 publications

(10 citation statements)

References 26 publications

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“…Portions are engulfed and sequestered as large vacuoles (56) in a sequence of events strikingly similar to that observed in embryogenesis of the stick insect Carausius morosus Br. (57). The B. germanica vacuoles then become vesiculated and partitioned into smaller vesicles (56) with the same size distribution as granules isolated from yolk (20).…”

Section: Discussionmentioning

confidence: 99%

A Cysteine Protease That Processes Insect Vitellin

Liu¹,

McCarron²,

Nordin

1996

Journal of Biological Chemistry

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A cysteine protease that initiates degradation of vitellin (Vt) in the orthopteran Blattella germanica, and its proprotease precursor, were purified from yolk and partially characterized. The protease, purified 300-fold, contains three peptides of M r 27,000, 29,000, and 31,000. A comparison of the purified enzyme's action pattern on Vt in vivo and in vitro confirmed its role in Vt processing. Protease-deficient yolk (day 0 postovulation) contained peptides of M r 35,500, 37,000, 39,000, and 41,000, which were absent from yolk with protease activity. These were replaced by three peptides of approximately M r 29,000, at days 2-3, the same time in development that protease expression and acidification of yolk granules occur (Nordin, J. H., Beaudoin, E. L., and Liu, X. (1991) Arch. Insect Biochem. Physiol. 18, 177-192). Acidification of purified proprotease converted it to three peptides of approximately M r 29,000 with cysteine protease activity. This conversion also required participation of a cysteine protease. Activated proprotease had the same pH activity profile, susceptibility to inhibitors, and cathepsin classification (L) as the protease. These results indicate that the Vt-processing protease is derived from a proprotease, which is activated in vivo by a developmentally regulated decrease in intragranular pH.

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Section: Discussionmentioning

confidence: 99%

A Cysteine Protease That Processes Insect Vitellin

Liu¹,

McCarron²,

Nordin

1996

Journal of Biological Chemistry

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“…For scanning electron-microscopic observations, yolk sacs were dissected in phosphate-buffered saline (PBS) and fixed in Karnovsky's fixative as previously reported (Fausto et al 1994). They were then thoroughly rinsed in 0.1 M cacodylate buffer at pH 7.2, postfixed for 2 h in 1% osmium tetroxide in 0.1 M cacodylate buffer at pH 7.2, dehydrated in an ascending series of alcohols and eventually critical point dried in a Balzer's apparatus equipped with a liquid CO 2 inlet.…”

Section: Structural Observationsmentioning

confidence: 99%

“…We have studied yolk utilisation in embryos of the stick insect Carausius morosus and found that yolk granules in this species are reconstituted anew with the beginning of embryonic development (Fausto et al 1994). By this time, the fluid ooplasm is gradually transformed into a collection of progressively smaller yolk granules due to invasion of secondary vitellophages from the egg periphery (Fausto et al 1997).…”

Section: Introductionmentioning

confidence: 99%

Yolk granules are differentially acidified during embryo development in the stick insect Carausius morosus

Fausto

Gambellini

Mazzini

et al. 2001

Cell and Tissue Research

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Newly laid eggs of stick insects comprise a unique fluid ooplasm that is gradually partitioned into a number of yolk granules by invasion of secondary vitellophages. This study aimed at establishing how yolk granules become acidified in the course of embryonic development. Data show that acidified yolk granules are rather scarce and randomly distributed in vitellophages of early embryos, while they tend to increase gradually in number as development proceeds to completion. Yolk granule acidification is progressively more inhibited in the presence of increasing concentrations of chloroquine, monensin and bafilomycin. A pro-protease was identified cytochemically and by immunoblotting in yolk extracts of progressively more advanced embryos. A specific monoclonal antibody raised against this pro-protease helped to demonstrate that it is gradually processed to yield a lower molecular weight polypeptide as development proceeds to completion. This latter polypeptide was identified as a protease using electrophoresis in polyacrylamide gels containing yolk extracts. Simultaneous administration of a fluorescent substrate for cysteine protease and an acidotropic probe produced superimposable labelling patterns, suggesting that only acidified yolk granules possess a proteolytic activity. On the other hand, yolk granules probed simultaneously for acidification and latent pro-protease yielded labelling patterns partially superimposed. Pro-protease labelling is gradually lost as yolk granules are progressively more acidified during development. Distinct labelling patterns were also obtained in vitellophages processed for the simultaneous detection of pro-protease and protease, suggesting that the two activities are expressed by different yolk granule populations, and that one is gradually converted into the other as time goes by.

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“…Yolk sacs were fixed in Karnovsky's fixative as previously reported (Fausto et al 1994). They were thoroughly rinsed in 0.1 M cacodylate buffer at pH 7.2, postfixed for an additional 2 h in 1% osmium tetroxide in 0.1 M cacodylate buffer at pH 7.2, dehydrated in an ascending series of alcohols, embedded in Epon-Araldite and allowed to polymerize for 3 days at 60°C.…”

Section: Structural Observationsmentioning

confidence: 99%

“…In hemimetabolous insects, embryos develop from a ventral thickening of the peripheral blastoderm, the so-called germ band, while the extra-embryonic blastoderm differentiates into such membranous envelopes as the inner amnion and the outer serosa membrane (Campos-Ortega & Hartenstein 1985;Pennacchio et al 1994). Newly laid eggs of the stick insect Carausius morosus are characterized by a yolk fluid ooplasm, which is partitioned into a number of yolk granules on invasion of secondary vitellophages from the egg periphery (Fausto et al 1994). A morphologically similar process appears to occur in other orthopteroid species (Petavy 1985;Giorgi & Nordin 1994).…”

Section: Introductionmentioning

confidence: 99%

Serosa membrane plays a key role in transferring vitellin polypeptides to the perivitelline fluid in insect embryos

et al. 2001

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In mid‐embryogenesis, the stick insect Carausius morosus comes to be comprised of three distinct districts: the embryo proper, the yolk sac and the perivitelline fluid. A monolayered epithelium, the so‐called serosa membrane, encloses the yolk sac and its content of vitellophages and large yolk granules. During embryonic development, the yolk sac declines gradually in protein concentration due to Vt polypeptides undergoing limited proteolysis to yield a number of Vt cleavage products of lower molecular weights. mAbs 1D1 and 5H11 are monoclonal antibodies raised against some of the Vt cleavage products generated by this process in the yolk sac. At the confocal microscope, antibody fluorescence is initially associated with a few yolk granules, while it is gradually displaced in the cytosolic spaces of the vitellophages. With the proceeding of embryonic development, label appears also in the serosa membrane in the form of clustered dots. At the ultrastructural level, gold particles are initially associated with the vitellophages that are labeled on a few yolk granules and in the cytosolic space flanking the yolk granules. Subsequently, the serosa cells become labeled on vesicles close to the yolk granules or just underneath the plasma membrane. Inside the serosa cells, label is also associated with granules budding from the Golgi apparatus, but never with the intercellular channels percolating the serosa membrane. These observations are interpreted as indicating that Vt cleavage products leak out from the yolk granules into the cytosolic spaces of the vitellophages and are eventually transferred to the perivitelline fluid via transcytosis through the serosa cells.

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An Ultrastructural Investigation on Vitellophage Invasion of the Yolk Mass during and after Germ Band Formation in Embryos of the Stick Insect Carausius morosus Br.

Cited by 16 publications

References 26 publications

A Cysteine Protease That Processes Insect Vitellin

A Cysteine Protease That Processes Insect Vitellin

Yolk granules are differentially acidified during embryo development in the stick insect Carausius morosus

Serosa membrane plays a key role in transferring vitellin polypeptides to the perivitelline fluid in insect embryos

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