Vision (Insects)

Vitellogenesis

Vitellogenesis is the process by which yolk accumulates in the cytoplasm of an ovarian oocyte. It is one of the final stages of egg formation, occurring just prior to deposition of the cho-rion. Studies on vitellogenesis have focused during the last 50 years on the major protein yolk precursor, vitellogenin, its synthesis in the fat body, its transport to the oocyte, its sequestration by receptor-mediated endocytosis, and the developmental and hormonal mechanisms that control these processes.

VITELLOGENIN SECRETION BY

THE FAT BODY

Before vitellogenesis can begin, the female fat body must transform from a tissue that supports molting, metamorphosis, and intermolt metabolism to one that can secrete vitellogenin and other yolk precursors. When the fat body has reached a requisite stage of maturity and hormonal stimulation, vitellogenin genes are transcribed that encode a polypeptide chain whose molecular weight in many insects is over 200,000. During transport through the endoplasmic reticulum and Golgi bodies, this polypeptide is clipped by endoproteases that remove a secretory signal from its N-terminal end and divide it into shorter polypeptide subunits. The latter differ in number from one in some Hymenoptera to nine in some Hemimetabola. As it moves through the secretory pathway, the complex of subunits is conjugated at sequence-specific sites with high-mannose oligosaccharides, phosphate, and lipids.
Amino acid sequences determined for vitellogenins from at least seven orders of insects are sufficiently similar to indicate a common ancestry. They are members of a protein superfamily that also includes the yolk proteins of vertebrates and nematodes, lipid transport proteins in the blood of both vertebrates and insects, and receptor proteins involved in the endocytosis of lipoproteins.
The yolk proteins of cyclorrhaphan Diptera are exceptional in that their amino acid sequences resemble those of a family of digestive enzymes, the vertebrate lipases. They nevertheless behave like conventional vitellogenins in being synthesized and conjugated in the female fat body, and deposited in cytoplasmic vesicles by the oocyte after endocytosis. A notable difference from conventional vitellogenins is that the several yolk polypeptides in a species are encoded by separate genes, rather than being proteolytic fragments of one gene product.
Other fat body products may supplement vitellogenin in the yolk. The eggs of several Lepidoptera contain lipophorin, a hemolymph protein whose functions include the delivery of lipids from the fat body to other tissues. In the yellow fever mosquito, Aedes aegypti, two proproteases are synthesized in the fat body and deposited in the eggs, where they are converted to active proteases during embryo-genesis. Endocytosis of hemolymph proteins that bind iron, calcium, heme groups, or biliverdin concentrates these ligands in the yolk of select species. Some of these supplementary proteins are secreted in synchrony with and under the same hormonal control as vitello-genin. Others, like lipophorin, serve somatic functions that require their presence in the hemolymph of males as well.


VITELLOGENIC FUNCTIONS IN OVARIAN FOLLICLES

In the ovaries, the morphological unit of vitellogenesis is a follicle—a single oocyte surrounded by an epithelium of somatic cells (the follicle cells) and associated in many insects with a set of modified germ cells (the nurse cells) (Fig. 1). All three cell types are necessary for vitellogenesis, but they contribute to it in very different ways.
Follicles begin to form during or shortly after metamorphosis. They are produced in linear chains termed ovarioles. Within each ovariole is a developmental gradient of follicles (Fig. 1), with the most mature one lying close to the beginning of the oviduct. A common pattern among cyclic egg producers is for only one follicle at a time in each ovariole to form yolk. The penultimate follicle is retarded in its development until the next reproductive cycle. In noncyclic insects, many follicles in each ovariole may simultaneously form yolk (Fig. 1).
Reflected light micrograph of a vitellogenic ovariole from the saturniid moth H. cecropia. The largest 36 follicles are vitellogenic. The yellow mass in each follicle is a yolk-filled oocyte, whose opacity is due to light scattering by yolk particles. The yellow color is due to carotenoids carried by vitellogenin and lipophorin, the two most abundant yolk proteins in saturniids. Nurse cells form a transparent cap at one end of each follicle. The epithelium of follicle cells surrounding each oocyte and its nurse cells is too thin to be readily visible, except where it forms a connection between successive follicles. Scale: largest follicle is about 1.9 mm long.
FIGURE 1 Reflected light micrograph of a vitellogenic ovariole from the saturniid moth H. cecropia. The largest 36 follicles are vitellogenic. The yellow mass in each follicle is a yolk-filled oocyte, whose opacity is due to light scattering by yolk particles. The yellow color is due to carotenoids carried by vitellogenin and lipophorin, the two most abundant yolk proteins in saturniids. Nurse cells form a transparent cap at one end of each follicle. The epithelium of follicle cells surrounding each oocyte and its nurse cells is too thin to be readily visible, except where it forms a connection between successive follicles. Scale: largest follicle is about 1.9 mm long.

Follicle Cells, Patency, and Secondary Yolk Proteins

At the onset of vitellogenesis, the follicle cells develop a system of intercellular spaces that give proteins from the hemolymph access to the surface of the oocyte. In the bug Rhodnius prolixus, patency has been attributed to cellular protrusions whose cytoskeletal elongation pushes neighboring follicle cells apart. In the cecropia moth, Hyalophora cecropia, osmotic shrinkage of the follicle cells is crucial. Whichever mechanism applies, the intercellular spaces are under tight developmental control: they arise at the onset of vitellogenesis and close when it terminates.
In addition, the follicle cells of many insects secrete proteins that are endocytosed by the oocyte along with vitellogenin. These products may resemble vitellogenin in size, antigenicity, and amino acid sequence; examples occur in Thysanura, Heteroptera, and Coleoptera. A similar relationship holds for the yolk proteins of the cyclorrhaphan Diptera. In Lepidoptera, follicle cell products are instead lipaselike sequences reminiscent of the yolk proteins of the Cyclorrhapha. A few exceptions are known in which either fat body or follicle cells but not both secrete precursors for the protein yolk.
Finally, the follicle cells connect to the oocyte during vitello-genesis via gap junctions that permit direct cytoplasm-to-cytoplasm transfer of ion currents and small organic molecules. These junctions have the potential to function in intercellular exchange of signaling substances such as cyclic nucleotides and calcium ions.

Nurse Cells and the Origin of Egg Cytoplasm

In all holometabolous and a few hemimetabolous orders, the oocytes connect via cytoplasmic bridges to nurse cells (Fig. 1). The bridges are wide enough to permit passage of ribosomes and membranous organelles such as mitochondria. Once believed to be the site of yolk production, nurse cells are now known to be the primary source of egg cytoplasm. They support vitellogenesis by providing the oocyte with the ribosomes, transcripts, and metabolic machinery needed to synthesize the receptors, structural proteins, and enzymes necessary for yolk deposition. In Orthoptera, Blattodea, and other Hemimetabola that lack nurse cells, the requisite transcripts are produced instead within the oocyte’s own nucleus by amplified nucleoli and lampbrush chromosomes.

Receptor-Mediated Endocytosis in the Oocyte

The surface of the vitellogenic oocyte contains receptors that can selectively bind vitellogenin and other yolk precursors after they have penetrated the spaces between the follicle cells. The receptors form transmembrane associations with clathrin lattices on the cytoplasmic side of the membrane. The membranes containing these complexes then fold inward to form endocytotic vesicles (Fig. 2 ). In subsequent processing steps, vitellogenin dissociates from its receptors within the vesicles, and the clathrin is released from the lattices on the outside. The denuded vesicles transfer their cargo of yolk precursors to neighboring yolk bodies by membrane fusion. Receptors, clath-rin lattices, and extra membrane are recycled to the oocyte surface for more rounds of endocytosis. In some insects, vitellogenin is later modified in the yolk to a less soluble storage form known as vitellin.
Electron micrograph of several endocytotic vesicles near the surface of a vitellogenic oocyte from H. cecropia. The bristlelike outer coat is the clathrin lattice that generates the force used to bend coated surface membranes into cytoplasmic vesicles. Receptor-bound vitellogenin is included in the thick layer of granular material that lines each vesicle. An incipient vesicle on the upper right is still attached to an infolding of the cell membrane. The average diameter of coated endocytotic vesicles is about 0.15 mm.
FIGURE 2 Electron micrograph of several endocytotic vesicles near the surface of a vitellogenic oocyte from H. cecropia. The bristlelike outer coat is the clathrin lattice that generates the force used to bend coated surface membranes into cytoplasmic vesicles. Receptor-bound vitellogenin is included in the thick layer of granular material that lines each vesicle. An incipient vesicle on the upper right is still attached to an infolding of the cell membrane. The average diameter of coated endocytotic vesicles is about 0.15 mm.
In the other two classes of yolk particles, glycogen is synthesized inside the oocyte from hemolymph-derived sugars, whereas lipid droplets are assembled from precursors carried from the fat body to the oocyte by lipophorin. Because the lipid droplets contain primarily triacylglycerols and lipophorin transports primarily diacylglycer-ols, enzymatic conversions must take place during the transfer.

CONTROL BY JUVENILE HORMONE AND ECDYSONE

In many insects, juvenile hormone secreted by the corpora allata stimulates the fat body of adult females to initiate the synthesis of vitellogenin. It may also promote patency of the follicle cells, hence making vitellogenin in the hemolymph available to the oocyte for endocytosis. Juvenile hormone, whose power as an inhibitor of metamorphosis requires that its secretion be reduced during pupation and adult development, has thus evolved new kinds of role in adults: it has become an effector of the neuroendocrine networks that synchronize vitellogenesis with feeding, the photoperiod, and mating.
Variations occur on this theme. In species such as stick insects, and native and domestic species of silk moths that complete egg formation during adult development, juvenile hormone is not required for vitellogenesis. In its classical role as an inhibitor of metamorphosis, the hormone may even prevent the fat body and ovaries from completing their essential previtellogenic development.
Another kind of variation occurs among Diptera, whose yolk protein synthesis by fat body is triggered by ecdysone. Juvenile hormone may still be required, but here it promotes posteclosion development of the fat body to a stage capable of responding to ecdysone. In the yellow fever mosquito, the ovaries themselves were shown to secrete ecdysone in response to EDNH (egg development neurosecre-tory hormone), a brain hormone released from the corpora cardiaca following a blood meal.

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