Studies on binding and uptake of vitellogenin by follicles of the tobacco hornworm, Manduca sexta

Abstract

AbstractAn in vitro system for the uptake of 125l‐vitellogenin (VG) or vitellin into isolated follicles of the tobacco hornworm, Manduca sexta, is described. After incubation with 125l‐VG, follicles were disrupted and the internal yolk contents separated from the follicle membranes. The results showed that 125l‐VG was associated principally with the membranes (92%) after incubation at 4°C. However, at 27°C, 125l‐VG was mainly in the yolk (92%). Furthermore, trypsin treatment removed approximately 70% of VG bound to the follicles at 4°C. Labeled VG was shown to bind to sonicated follicle membranes with high specificity and affinity (KD ⋍ 1.3 × 10−8 M). This binding was sensitive to pH and calcium concentration. The total binding sites were estimated at 4 × 1014 sites/g of membrane protein. Competition studies showed that binding of 125l‐VG to follicle membranes was blocked by excess unlabeled vitellin and deglycosylated vitellogenin but not by lipophorin (the major hemolymph lipoprotein), microvitellogenin, a female‐specific protein (Mr ∼ 31,000) found in both hemolymph and eggs, and the smaller vitellogenin subunit, apovitellogenin‐II (Mr ∼ 45,000). These results suggest that selective uptake of M. sexta VG from the hemolymph involves binding to specific receptors located on the follicle membranes.