Vitellogenin Subunits Research Articles

The N-terminal subunit of vitellogenin in planthopper eggs and saliva acts as a reliable elicitor that induces defenses in rice.

Vitellogenins (Vgs) are critical for the development and fecundity of insects. As such, these essential proteins can be used by plants to reliably sense the presence of insects. We addressed this with a combination of molecular and chemical analyses, genetic transformation, bioactivity tests, and insect performance assays. The small N-terminal subunit of Vgs of the planthopper Nilaparvata lugens (NlVgN) was found to trigger strong defense responses in rice when it enters the plants during feeding or oviposition by the insect. The defenses induced by NlVgN not only decreased the hatching rate of N.lugens eggs, but also induced volatile emissions in plants, which rendered them attractive to a common egg parasitoid. VgN of other planthoppers triggered the same defenses in rice. We further show that VgN deposited during planthopper feeding compared with during oviposition induces a somewhat different response, probably to target the appropriate developmental stage of the insect. We also confirm that NlVgN is essential for planthopper growth, development, and fecundity. This study demonstrates that VgN in planthopper eggs and saliva acts as a reliable and unavoidable elicitor of plant defenses. Its importance for insect performance precludes evolutionary adaptions to prevent detection by rice plants.

Identification of Receptor‐Interacting Regions of Vitellogenin within Evolutionarily Conserved β‐Sheet Structures by Using a Peptide Array

Vitellogenesis, a key process in oviparous animals, is characterized by enhanced synthesis of the lipoprotein vitellogenin, which serves as the major yolk-protein precursor. In most oviparous animals, and specifically in crustaceans, vitellogenin is mainly synthesized in the hepatopancreas, secreted to the hemolymph, and taken up into the ovary by receptor-mediated endocytosis. In the present study, localization of the vitellogenin receptor and its interaction with vitellogenin were investigated in the freshwater prawn Macrobrachium rosenbergii. The receptor was immuno-histochemically localized to the cell periphery and around yolk vesicles. A receptor blot assay revealed that the vitellogenin receptor interacts with most known vitellogenin subunits, the most prominent being the 79 kDa subunit. The receptor was, moreover, able to interact with trypsin-digested vitellogenin peptides. By combining a novel peptide-array approach with tandem mass spectrometry, eleven vitellogenin-derived peptides that interacted with the receptor were identified. A 3D model of vitellogenin indicated that four of the identified peptides are N-terminally localized. One of the peptides is homologous to the receptor-recognized site of vertebrate vitellogenin, and assumes a conserved β-sheet structure. These findings suggest that this specific β-sheet region in the vitellogenin N-terminal lipoprotein domain is the receptor-interacting site, with the rest of the protein serving to enhance affinity for the receptor. The conservation of the receptor recognition site in invertebrate and vertebrate vitellogenin might have vast implications for oviparous species reproduction, development, immunity, and pest management.

Two-dimensional proteomic analysis of honeybee, Apis mellifera, winter worker hemolymph

Honeybee (Apis mellifera Linnaeus) colonies in temperate zones produce either summer bees, which have a lifespan of 15 to 48 days, or winter bees, which emerge in late summer and live up to 8 months. Winter bees develop unique physiological conditions characterized by changes in protein composition that appear to be major determinants of honeybee lifespan. We analyzed winter honeybee worker hemolymph using a proteomic approach for the first time. Hemolymph collected from the dorsal vessel of winter honeybees using a glass capillary tube was analyzed using two-dimensional gel electrophoresis followed by MALDI TOF/TOF protein identification. Overall, 93 spots were assigned significance (P < 0.05). Many identified proteins corresponded well with extended lifespan. Vitellogenin subunits (mainly ∼180 and ∼100 kDa) comprised the major portion of the proteins; however, vitellogenin dominance repressed the signals of the lower-abundance proteins. Future physiological studies related to overwintering bees, including health, immunity, longevity, nutrition, and/or colony losses, can benefit from these results.

Estrogenic effects of monocrotophos evaluated by vitellogenin mRNA and protein induction in male goldfish ( Carassius auratus)

Monocrotophos (MCP) is listed as a Prior Informed Consent chemical, and it is still used extensively in developing countries. It has been seen that MCP shows high toxicity in fish and aquatic ecosystems. To confirm whether MCP shows estrogenic effects on fish, the induction of vitellogenin (VTG) in male goldfish ( Carassius auratus) following a 21 day exposure to 0.01, 0.10 and 1.00 mg·L − 1 40% MCP pesticide in a semi-static exposure system was evaluated by hepatic mRNA and plasma protein. Real-time reverse transcriptase polymerase chain reaction assay showed that MCP significantly increased hepatic mRNA expression of VTG in male goldfish in a dose-dependent manner. Western blot analysis revealed the presence of two VTG subunits with molecular masses of 134 kDa and 110 kDa in the plasma of the exposure groups, which were not observed in that of the control males, indicating the induction of VTG protein synthesis and secretion by MCP in male goldfish. Furthermore, the levels of plasma VTG protein in the control group and the 0.01, 0.10 and 1.00 mg·L − 1 40% MCP pesticide exposure groups were quantified by enzyme-linked immunoassay, and revealed concentrations of 53.8 ± 5.6 ng·mL − 1 , 285.5 ± 5.0 ng·mL − 1 , 972.9 ± 7.4 ng·mL − 1 and 678.4 ± 38.0 ng·mL − 1 , respectively. The overlapping results of VTG mRNA and protein indicated that MCP had significant estrogenic properties and was thus a potential endocrine disruptor.

Vitellogenesis in the sand shrimp, metapenaeus ensis: the contribution from the hepatopancreas-specific vitellogenin gene (MeVg2).

An additional vitellogenin gene (MeVg2) that is structurally different from MeVg1 was cloned and characterized from the shrimp Metapenaeus ensis. The MeVg2 gene consists of fewer exons-introns and is most likely evolved from the MeVg1 gene. The cDNA for MeVg2 is 8.0 kilobases (kb) in size, and the deduced MeVg2 precursor shared an overall 54% amino sequence identity to the MeVg1 gene of the same shrimp. As compared to the MeVg1 precursor, MeVg2 precursor consists of more potential subunit cleavage sites, suggesting that the precursor may be processed into many smaller subunits. The MeVg2 is expressed only in the hepatopancreas, and the expression level of MeVg2 in adult female increases from the early vitellogenic stage, reaching a maximum at the middle vitellogenic stage, and remains high toward the end of vitellogenic cycle. In addition to the 8-kb mRNA, smaller transcripts of 1.5-2.5 kb for MeVg2 were identified, and the 8-kb transcript only constitutes less than 10% of the overall MeVg2-derived transcripts. To confirm the presence of the small transcripts, we screened the shrimp hepatopancreas cDNA library and isolated two smaller MeVg2-specific cDNA clones. These clones shared greater than 99% overall identity to the corresponding C-terminal region of the MeVg2 precursor, suggesting that an alternative expression/ splicing of the MeVg2 gene occurred. By immunohistochemical analysis, vitellin-immunopositive signals were localized in the lumen and extracellular fraction of the hepatopancreas. Amino acid sequence determination of the tissue protein and secreted protein from the hepatopancreas revealed that the 76-kDa vitellogenin subunit is most likely processed into smaller-sized subunits. Taken together, these results suggest that the hepatopancreas is an important organ for the synthesis of vitellogenin and may contribute to vitellogenesis by producing a large quantity of smaller MeVg2 subunit for ovarian uptake.

Organization of the shrimp vitellogenin gene: evidence of multiple genes and tissue specific expression by the ovary and hepatopancreas

Vitellogenin is the major egg yolk protein synthesized in female shrimp during gonad maturation. Although there are several reports for the cloning of vitellogenin complementary DNA (cDNA) in different crustaceans, little is known of the gene organization of this protein. This study reports the first cloning and characterization of a full-length gene encoding the vitellogenin precursor from the shrimp Metapenaeus ensis. By genomic DNA library screening, six different lambda clones were isolated using shrimp partial gene sequence as probe. Initial DNA sequence determination revealed that these clones are derived from different genes with coding sequence similar to other crustacean vitellogenins. Two of these clones were used for further analysis. One of the lambda clones (λ3.3) carries most of the coding sequence that correspond to the M. ensis vitellogenin gene (MeVg1) and the other clone (λ8.3) carries a smaller portion of the coding sequence of a different vitellogenin gene (MeVg2). The λ3.3 clone was chosen for further characterization. To clone the remaining 5′ end upstream promoter region, 5′ untranslated region and the remaining coding sequence of MeVg1, a polymerase chain reaction (PCR)-based gene walking approach was used. Subsequently, a PCR clone with overlapping sequence identical to the genomic clone was obtained and the organization of MeVg1 gene was constructed. The MeVg1 gene consists of 15 exons and 14 introns spanning approximately 10 kb. Several potential cleavage sites were identified from the deduced vitellogenin precursor. Cleaving of the precursor in these sites would result in the production of several vitellogenin subunits. To clone the cDNA for the vitellogenin, 5′ and 3′ rapid amplification of cDNA ends was performed using ovary cDNA of the shrimp. A 4.4 kb 5′ cDNA clone and a 4 kb 3′ end cDNA clone were isolated. The size of the reconstructed cDNA for M. ensis Vg is 7.97 kb and consists of the longest open reading frame of 7776 bp. Unlike the vitellogenin precursor of most insects and vertebrates, the deduced vitellogenin precursor lacks the polyserine domain important for receptor-mediated endocytosis. Phylogenetic analysis revealed a closer relationship of the MeVg1 with other crustacean vitellogenins but distantly related to other invertebrate and vertebrate vitellogenins. By reverse transcription-PCR, we have demonstrated that the shrimp MeVg1 gene is expressed only in the ovary and hepatopancreas while the MeVg2 gene is expressed exclusively in the hepatopancreas. In conclusion, the shrimp ovary also contribute significantly in the production of vitellogenin at transcription level and the gene organization of the shrimp protein may provide an insight in the evolution of this group of important proteins.

Hepatopancreas is the likely organ of vitellogenin synthesis in the freshwater prawn, Macrobrachium rosenbergii.

The objective of the present study was to investigate the source of vitellogenin in the freshwater prawn, Macrobrachium rosenbergii. Ovarian development of M. rosenbergii was classified into five stages (stage I-V). Vitellin/vitellogenin was detected in the ovary and the hepatopancreas in different stages by native-PAGE and Western blotting. Two and three subunits of vitellin were observed in the ovary at the early- (I-II), mid- and late- (III-V) stages, respectively. The subunit of vitellogenin was not detected in the hepatopancreas at different stages of prawns. Hepatopancreas had positive immunocytological staining (against vitellin antibody) in different ovarian stages of prawn. Only vitellogenic oocyte but not previtellogenic oocytes and follicle cells had a positive immunocytological staining. Hepatopancreas could synthesize radiolabeled immunoreactive proteins after incubation with radiolabeled glycine on the basis of immunoprecipitation (against vitellin antiserum). Therefore, it is concluded that hepatopancreas is the most likely organ to synthesize vitellogenin in the freshwater prawn, M. rosenbergii.

Cyclic nucleotide-dependent protein phosphorylation in vitellogenic follicles of Hyalophora cecropia

(1) In homogenates of vitellogenic follicles from Hyalophora cecropia, cyclic nucleotides promoted the transfer of label from [γ 32P]-ATP to at least four polypeptides. PKI (6–20) amide, an inhibitor of PKA (cAMP-dependent protein kinase), prevented all four reactions. Quantitative tests using kemptide as a substrate indicated that 80% of the total follicular PKA activity was localized in the follicle cells; labeling at 45, 32, and 27 kDa was particle-associated and also restricted to the follicle cells, while a 58 kDa substrate was labeled only in homogenates of the oocyte. (2) When intact follicles were incubated in [ 32P]-phosphate and okadaic acid, a protein phosphatase inhibitor, the 32 kDa substrate again exhibited cAMP-dependent labeling. There was thus a physiological relationship between PKA activation and 32 kDa protein phosphorylation, while exposure of at least two of the other three substrates to appropriate kinases required homogenization. The latter was illustrated by phosphorylation of the 42 kDa small subunit of vitellogenin, which occurred only when homogenization mixed the proteins of the yolk bodies with cytoplasmic kinases. (3) PKA activation is known to promote the termination of vitellogenesis, even in the absence of detectable labeling of the 32 kDa substrate. The possibility remains that phosphorylation at 32 kDa concerns later aspects of postvellogenic development that were not tested by the assay system used here.

Vitellogenin and yolk protein processing in Bothrops jararaca (Wied), a tropical venomous snake

The plasma vitellogenin of Bothrops jararaca is composed of two subunits. The larger subunit (160 kDa) is phosphate rich and carbohydrate poor, while the smaller (110 kDa) is highly glycosylated and less phosphorylated. As in other vertebrates, the vitellogenin of B. jararaca is synthesized in the liver under estrogen control. The newly synthesized vitellogenin molecule is a 270 kDa polypeptide. This polypeptide originates the two subunits of the plasma vitellogenin by proteolytic cleavage. In the eggs of B. jararaca six main yolk polypeptides have been detected (113, 107, 104, 72, 27.2 and 20.7 kDa). Using phosphoprotein staining we have shown that the 72 kDa polypeptide is the larger phosvitin so far described in a vertebrate egg yolk. The 107 kDa yolk polypeptide also seems to be phosphorylated, but to a lesser extent than the phosvitin. The 104 kDa vitellin originates from the larger vitellogenin subunit while the 113 kDa vitellin originates from the smaller vitellogenin subunit. Based on these results we propose a general scheme for vitellogenin and vitellin processing in B. jararaca.

Localization of vitellogenin production in the blacklegged tick, Ixodes scapularis (Acari: Ixodidae)

Summary Vitellogenin synthesis was investigated in the ixodid tick, Ixodes scapularis. The fat body was the most active tissue in vitellogenin production. Midgut and ovarian tissues showed slight activity (~2000 cpm) which may have been due to a small amount of contamination by fat body cells. Proteins synthesized by the fat body were examined and confirmed as vitellogenin by electrophoresis and audioradiography. There were seven radioactive polypeptides corresponding to the subunits of vitellogenin. Immunochemical analysis showed a strong reaction with anti-vitellin antibody in the fat body and developing ovaries; however, no specific fluorescence was seen in the midgut. These results indicate that the fat body is the main and probably the only site of vitellogenin synthesis in this prostriate tick.

A18 トウゴウヤブカ小分子ビテロジェニン cDNA のクローニング

A18 トウゴウヤブカ小分子ビテロジェニン cDNA のクローニング

Vitellogenesis in virgin and mated females of the mealworm beetle, Tenebrio molitor.

A comparison between vitellogenesis in virgin and mated females of Tenebrio molitor showed significant differences at each investigated developmental stage. Yolk protein deposition in oocytes, measured as an increase in their size parameters (length, width, and volume), proceeded much faster and was more efficient in mated females as compared to virgins. In fertilized females the gonadotropic cycle showed a cyclicity with an eight-day period while virgin females finish their vitellogenic stage after the first cycle. These differences were reflected in changes in the rate of protein synthesis in the fat body of females completing vitellogenesis or entering the next oogenetic cycle. In the haemolymph, in addition to a large (158 kDa) and two small (56 kDa and 45 kDa) subunits of vitellogenin, there was an abundance of proteins of 80 kDa and 60 kDa.

CDNA cloning and expression of the large subunit of juvenile hormone-regulated vitellogenin from Lymantria dispar

The 5.4 kb RNA enriched in mRNA coding for the large subunits of vitellogenin (VgL mRNA) in the gypsy moth ( Lymantria dispar) was purified, and used to construct a cDNA library. One of the clones, pVL-80, had sequences representing an internal region of VgL mRNA. The cDNA representing the 5′-terminal region of VgL mRNA was isolated by 5′-RACE (Rapid Amplification of cDNA Ends), and found to contain sequences encoding the N-terminal amino acid sequence of Vg 190. The insert in pVL-80 was 822 bp long and had an open reading frame (ORF) extending beyond the insert in both directions. In vitro expression of the ORF produced a 32 kDa protein which reacted with Vg 190 antiserum. Southern analysis of gypsy moth genomic DNA suggested that a single copy of the VgL gene is present per genome. The pVL-80 was demonstrated to be useful as a specific and sensitive probe to investigate developmental and juvenile hormone regulation of VgL mRNA in the fat body.

Structure and expression of mRNA for vitellogenin in Bombyx mori

Vitellogenin, a precursor of major yolk protein of the silkworm, Bombyx mori is a tetramer composed of each two molecules of heavy and light subunits. We cloned mRNA sequence for the B. mori vitellogenin and analyzed its structure. Sequence alignment of several overlapping cDNA clones indicated that the vitellogenin mRNA is approx. 5.7 kb, containing an open reading frame for a peptide with 1782 amino acid residues. By comparing the deduced amino acid sequence with the amino-terminal primary structures of vitellogenin subunits, it is suggested that the heavy and light subunits of the B. mori vitellogenin are encoded by a single contiguous mRNA. The primary translation product of the vitellogenin mRNA was detected in the microsomal fraction prepared from the fat body of vitellogenic females. Northern blot analysis of the fat body RNA demonstrated that the biosynthesis of vitellogenin in B. mori is regulated in a tissue-, sex- and stage-specific manner at the level of mRNA. Possible cause for discrepancy between the present results and our previous proposal (Izumi, S. and Tomino, S. (1983) Insect Biochem. 13, 81–85) on the biosynthesis of B. mori vitellogenin is also discussed.

Analysis of Mosquito Vitellogenin cDNA: Similarity with Vertebrate Phosvitins and Arthropod Serum Proteins

The cDNA coding for vitellogenin of the mosquito Aedes aegypti was cloned and sequenced. An immunological analysis of expressed deletions from the 5′-end of the vitellogenin cDNA clones using vitellogenin subunit-specific antibodies showed that the small vitellogenin subunit is located at the N terminus and the large one at the carboxy-portion of the pre-pro-vitellogenin. The position of the cleavage between the vitellogenin subunits in the pre-pro-vitellogenin was identified by locating the N terminus of the large subunit. The cleavage site has a consensus RXRR for the subtilisin-processing endoprotease. Mosquito vitellogenin is highly hydrophilic with 17 putative N-linked glycosylation sites and 13 potential tyrosine sulfation sites. In contrast to known invertebrate vitellogenins, mosquito vitellogenin contains three polyserine domains that are similar to those of phosvitins in vertebrate vitellogenins. These polyserine domains, originally presumed to be vertebrate-specific, have several phosphorylation consensus sites in their sequences. Unlike other known vitellogenins, mosquito vitellogenin is rich in aromatic amino acid residues, tyrosine and phenylalanine, and in this respect is similar to insect serum proteins, arylphorins. This similarity suggests that mosquito vitellogenin may supply aromatic amino acids to the cuticle of rapidly developing embryos.

Juvenile hormone regulation of vitellogenin in the gypsy moth, Lymantria dispar: Suppression of vitellogenin mRNA in the fat body

Hormonal regulation of vitellogenin synthesis and accumulation in the gypsy moth, Lymantria dispar, was examined using a potent juvenile hormone analogue fenoxycarb. The vitellogenin in the haemolymph of female larvae could be detected on days 4–5 of the 5th (final) larval stadium and continued to accumulate throughout the remainder of the stadium. Treatment of larvae with fenoxycarb during the intermoult period from 4th to 5th instar caused a delay in the appearance of the vitellogenin in the haemolymph and the rate of vitellogenin accumulation decreased. This effect of fenoxycarb was dose responsive and depended on the developmental stage of the larva. Treatment done after day 2 of the 5th stadium had no effect on the vitellogenin accumulation. Analysis of poly(A) RNA isolated from the fat body of female larvae showed the presence of a 5400 nucleotide (nt) RNA species in untreated larvae not present in fenoxycarb-treated larvae. This RNA species was not detected in the fat body of day 7 5th-instar male and day 1 5th-instar female larvae but was present in females on day 6 and later part of the 5th stadium. Translation of poly(A) RNA from untreated female larvae yielded a protein of 175 kDa whereas poly(A) RNA from fenoxycarb-treated female larvae did not. The presence of the 175 kDa product depended upon the presence of the 5400 nt RNA species in the poly(A) RNA. The 175 kDa translation product was immunologically related to the large subunits of the vitellogenin. A synthetic oligonucleotide probe encoding the amino acid sequence corresponding to the N-terminal region of the large subunit of vitellogenin (Vg190) hybridized specifically to the 5400 nt RNA. These data indicate that fenoxycarb acts by regulating the level of vitellogenin mRNA in the fat body.

Temporal events of gypsy moth vitellogenesis and ovarian development

Abstract The vitellogenic period of gypsy moth ovarian development starts on day 3 of the pupal stage and continues through adulthood. During this period, rapid increases occur in follicle size, protein content, and wet weight of the ovary. Patency is observed on day 3 of the pupal stage.Pre‐vitellogenic follicles are formed in the last larval stadium. Newly formed follicles detach from the germarium on day 4, and increase rapidly to 140 per ovariole at the end of the last larval stadium. The pre‐vitellogenic follicles are uniformly around 50 um in diameter. No vitellogenin is incorporated into the oocytes until the pupal stage.Polyacrylamide gel electrophosesis (PAGE) in the presence of sodium dodecylsulphate (SDS) analysis of male and female haemolymph samples and vitellogenic ovaries demonstrates the presence of two female‐specific subunits of vitellogenin of 180 kD and 160 kD. These proteins are detected only in haemolymph and ovarian extracts of vitellogenic females. The molecular weight of the native protein determined by size exclusion chromatography is approximately 400–420 kD.A highly sensitive double antibody sandwich enzyme‐linked immunosorbent assay (ELISA) was developed to monitor the temporal changes in vitellogenin titre in haemolymph. Vitellogenin production starts on day 2 of the last larval stadium, reaching a maximum level by day 6 of the last larval stadium, and decreasing in the late pupal stage as vitellogenin was internalized into the oocytes. This is the first report of vitellogenin production occurring in the larval stage of a holometabolous insect. The fact that vitellogenin production and uptake occur during different stages of development in the gypsy moth, opens up some interesting questions concerning the underlying regulatory mechanisms controlling each process.

Mosquito vitellogenin subunits originate from a common precursor

Using a cell-free translation system, we demonstrated that the two subunits of mosquito vitellogenin (VG), 200 kDa and 65 kDa, originate from a common precursor. The precursor polypeptide of 220 kDa is a translation product specific to mRNA from vitellogenic mosquitoes. In immunoprecipitation analysis, the 220-kDa polypeptide was recognized by monoclonal antibodies directed either to the large or the small VG subunit. Peptide mapping showed homology between the 220-kDa polypeptide and both subunits, thus providing further proof that the 220-kDa product of translation is the precursor for both VG subunits. In the presence of microsomal membranes, the molecular size of the VG precursor increased to 235 kDa suggesting this as a first step in co-translational modifications of VG.

Properties of the mosquito yolk protein: A study using monoclonal antibodies

The yolk protein (YP) of the mosquito Aedes aegypti was analyzed with radiolabeling experiments and monoclonal antibodies (mABs). Immunoprecipitation analyses with mAB demonstrated that the mature vitellogenin (VG) secreted by the fat body consisted of two subunits, 200 and 65 kDa. In the oocyte, vitellin (VT) was also composed of only these two subunits, and no additional processing of VT was detected after its internalization and crystallization. Immunoprecipitation experiments showed that the YP subunits were not associated covalently, since they behaved similarly during SDS-PAGE under reduced or non-reduced conditions. Labeling with radioactive carbohydrate precursors revealed that both VG subunits were glycosylated and retained their carbohydrate moieties after their internalization and processing into VT by the oocyte. They were composed of mannose and N- acetylglucosamine . The carbohydrate moieties of VG from the fat body were sensitive to endo-β-N- acetylglucosaminidase H (Endo H), and they continued to be Endo H-sensitive in crystalline VT in the oocyte. Therefore, the carbohydrate moieties of both YP subunits were high-mannose oligosaccharides. Treatment of [ 35S]methionine labeled VG and VT by Endo H demonstrated that carbohydrate moieties accounted for 10 kDa in a large subunit and for 15 kDa in a small subunit. Labeling with [ 32P]orthophosphate showed that both subunits of VG and VT were phosphorylated, and phosphorylated moieties were resistant to delipidation.

Biosynthesis, purification, and characterization of Aedes aegypti vitellin and vitellogenin

AbstractVitellin, the major egg yolk protein, and vitellogenin, the hemolymph precursor of egg yolk protein, have been purified to apparent homogeneity from the mosquito Aedes aegypti. The purification procedure included chromatography on ion exchange, hydrophobic, and gel filtration columns. Vitellin and vitellogenin have a similar molecular weight (Mr 300,000) on gel filtration columns. However, the molecular weights of vitellin and vitellogenin, as determined from SDS electrophoresis, were 393,000 and 337,000, respectively.Vitellin in sodium dodecyl sulfate released six subunits of molecular weight 116,000, 83,000, 75,000, 54,000, 36,000, and 29,000, whereas vitellogenin released only three subunits (155,000, 120,000, and 62,000). The average molecular weights of vitellin and vitellogenin after gel filtration and SDS electrophoresis were 346,000 and 318,000, respectively.Vitellin has a high content of aspartic acid and glutamic acid, and a low content of histidine, methionine, cysteine, and tryptophan. Vitellin also contains 0.9% mol of glucosamine and no galactosamine. The isoelectric points of vitellin and vitellogenin are at pH 6.4 and 6.3, respectively.Aedes aegypti fat bodies incubated for short intervals in tissue culture medium in the presence of [3H]valine showed incorporation by radio‐immunoprecipitation and SDS electrophoresis into three primary vitellogenin polypeptides of molecular weights (± SEM) 156,000 ± 4,000, 114,000 ± 5,000, and 62,000 ± 400 inside the fat body and 162,000 ± 3,000, 118,200 ± 2,000, and 63,000 ± 300 in the medium. These results suggest that the molecular weight of vitellogenin synthesized inside the fat body (Mr 332,000) remains unchanged when secreted into the hemolymph (Mr 343,000). The three vitellogenin subunits are processed by the ovary into six subunits which are then deposited in the yolk granules as vitellin.