3 research outputs found
Polyphosphate polymers during early embryogenesis of Periplaneta americana
8 p. : il.Inorganic polyphosphates (PolyP) are linear polymers of phosphate (Pi) residues linked by high-energy phosphoanhydride bonds. Despite a wide distribution, their role during insect embryogenesis has not been examined so far. In this study, we show the mobilization of PolyP polymers during the
embryogenesis of the cockroach Periplaneta americana. PolyP was detected by enzymatic and fluorimetric assays and found to accumulate in two main sizes by agarose gel electrophoresis. Confocal microscopy showed their presence in small vesicles. In addition, X-ray microanalysis of small vesicles showed considerable amounts of calcium, sodium and magnesium, suggesting an association of PolyP with these elements. Variations of the free Ca+2, Pi and PolyP levels were observed during the first days of
embryogenesis. Our results are consistent with the hypothesis that phosphate ions modulate PolyP variation and that PolyP hydrolysis result in increasing free Ca+2 levels. This is the first investigation of
PolyP metabolism during embryogenesis of an insect and might shed light on the mechanisms involving Pi storage and homeostasis during this period. We suggest that PolyP, mainly stored in small vesicles,
might be involved in the functional control of Ca+2 and Pi homeostasis during early embryogenesis of P. Americana
Inorganic polyphosphate inhibits an aspartic protease-like activity in the eggs of rhodnius prolixus (Stahl) and impairs yolk mobilization in vitro
6 p. : il.Inorganic polyphosphate (poly P) is a polymer of phosphate residues that has been shown to act as modulator of some vertebrate
cathepsins. In the egg yolk granules of Rhodnius prolixus, a cathepsinDis the main protease involved in yolk mobilization and is dependent on an activation by acid phosphatases. In this study, we showed a possible role of poly P stored inside yolk granules on the inhibition of cathepsinDand arrest of yolk mobilization during early embryogenesis of these insects. Enzymatic assays detected poly P stores inside the
eggs of R. prolixus. We observed that micromolar poly P concentrations inhibited cathepsin D proteolytic activity using both synthetic peptides and homogenates of egg yolk as substrates. Poly P was a substrate for Rhodnius acid phosphatase and also a strong competitive inhibitor of a pNPPase activity. Fusion events have been suggested as important steps towards acid phosphatase transport to yolk granules.
Weobserved that poly P levels in those compartments were reduced after in vitro fusion assays and that the remaining poly P did not have the same cathepsinDinhibition activity after fusion. Our results are consistent with the hypothesis that poly P is a cathepsinDinhibitor and a substrate for acid phosphatase inside yolk granules. It is possible that, once activated, acid phosphatase might degrade poly P, allowing
cathepsin D to initiate yolk proteolysis. We, therefore, suggest that degradation of poly P might represent a new step toward yolk
mobilization during embryogenesis of R. prolixus
Proton-pyrophosphatase and polyphosphate in acidocalcisome-like vesicles from oocytes and eggs of periplaneta americana
9 p. : il.Acidocalcisomes are acidic organelles containing large amounts of polyphosphate (poly P), a number of
cations, and a variety of cation pumps in their limiting membrane. The vacuolar proton-pyrophosphatase
(V-HĂľ-PPase), a unique electrogenic proton-pump that couples pyrophosphate (PPi) hydrolysis to the
active transport of protons across membranes, is commonly present in membranes of acidocalcisomes. In
the course of insect oogenesis, a large amount of yolk protein is incorporated by the oocytes and stored in
organelles called yolk granules (YGs). During embryogenesis, the content of these granules is degraded
by acid hydrolases. These enzymes are activated by the acidification of the YG by a mechanism that is
mediated by proton-pumps present in their membranes. In this work, we describe an HĂľ-PPase activity
in membrane fractions of oocytes and eggs of the domestic cockroach Periplaneta americana. The enzyme
activity was optimum at pH around 7.0, and was dependent on Mg2Ăľ and inhibited by NaF, as well as by
IDP and Ca2Ăľ. Immunolocalization of the yolk preparation using antibodies against a conserved sequence
of V-HĂľ-PPases showed labeling of small vesicles, which also showed the presence of high concentrations
of phosphorus, calcium and other elements, as revealed by electron probe X-ray microanalysis. In
addition, poly P content was detected in ovaries and eggs and localized inside the yolk granules and the
small vesicles. Altogether, our results provide evidence that numerous small vesicles of the eggs of
P. americana present acidocalcisome-like characteristics. In addition, the possible role of these organelles
during embryogenesis of this insect is discussed