The Drosophila yolkless gene encodes a vitellogenin receptor belonging to the low density lipoprotein receptor superfamily.

Abstract

Sequence comparisons of vitellogenins from a wide range oforganisms have identified regions of similarity not only to each other but also to vertebrate apolipoproteins (e.g. apoB-100 and apoE). Furthermore, the chicken vitellogenin receptor, which also binds apolipoproteins, has been found to belong to the low density lipoprotein receptor (LDLR) superfamily [Bujo, H., Hermann, M., Kaderli, M. O., Jacobsen, L., Sugawara, S., Nimpf, J., Yamamoto, T. & Schneider, W. J. (1994) EMBO J. 13,[5165][5166][5167][5168][5169][5170][5171][5172][5173][5174][5175]. The yolk proteins of higher dipterans are exceptional, however, and instead show similarity to lipoprotein lipases. The molecular characterization of the putative Drosophila melknogaster vitellogenin receptor gene, yolkless (yl), described in this report reveals that the protein it encodes (Yl), is also a member of the LDLR superfamily. The ovary-specific 6.5-kbyl RNA codes for a protein of -210 kDa which contains all three motifs common to the LDLR class of proteins. Within this superfamily, Yl may be related more to the LDLR-related proteins (LRPs), which bind both apolipoproteins and lipoprotein lipases. The similarity of Y1 to the other LDLR proteins is restricted to the putative extracellular domain. Most noticeably, the cytoplasmic domain of Yl lacks the typical NPXY sequence which is involved in receptor internalization.One of the best-studied mechanisms for the selective uptake of proteins into cells is receptor-mediated endocytosis. Two key advances in the study of endocytosis have been the development of cell-free assays and the identification of mutations which disrupt specific steps of endocytosis (1, 2). In this regard, we have been interested in identifying mutations which affect endocytosis in Drosophila. We have focused on the uptake of vitellogenins (yolk proteins) into the developing egg for several reasons. First, oocytes incorporate large amounts of vitellogenins (3, 4). Second, in oocytes, a specialized mechanism has evolved so that the yolk proteins are stored in yolk granules instead of being fated for degradation in lysosomes. Third, in many insects, juvenile hormone is required for the oocyte to initiate vitellogenin uptake (4, 5).Obviously, one of the key components in the uptake of vitellogenins is the receptor, and although vitellogenins from a variety of organisms have been characterized, much less is known about their receptors. In vertebrates, one of the beststudied systems is the chicken oocyte 95-kDa receptor, which binds two components of chicken yolk-vitellogenin and very low density lipoproteins (6). Mutants lacking this receptor fail to accumulate yolk proteins in the oocyte, and plasma vitellogenin levels are elevated (6, 7). Recently, Bujo et al. (8) have shown that the 95-kDa receptor is closely related to the mammalian very low density lipoprotein receptor (VLDLR). Frog and fish vitellogenin receptors appear to be related to the chicken receptor (9, 10).By using ligand-binding assays, vitellogenin receptors of simil...

Regulation of the Vitellogenin Receptor duringDrosophila melanogasterOogenesis

“…Furthermore, they identified a minimal enhancer region sufficient for germ line-specific expression of yl. Previously, a genomic DNA fragment that contained the yl gene, including a region 1.7 kb upstream of the start site, was shown to be sufficient for rescue of the yl mutant phenotype (Schonbaum et al, 1995). We cloned portions of this 1.7-kb region into a reporter vector bearing an hsp70 basal promoter element linked to the lacZ gene (Thummel and Pirrotta, 1991).…”

“…DNAs were purified (Midi-Preps; Qiagen, Hilden, Germany) and coprecipitated with a helper P transposase plasmid (phs ). DNAs were injected into y w 1118 hosts and selected for white ϩ phenotype as described previously (Schonbaum et al, 1995). Transgenic lines were tested for ␤-galactosidase activity with 5-bromo-4-chloro-3-indolyl-␤-d-galactopyranoside (Margolis and Spradling, 1995).…”

“…Random-primed digoxygenin-labeled DNA probes were prepared as described by the manufacturer (Boehringer Mannheim, Indianapolis, IN). The yl cDNAs used for the in situ hybridizations have been described previously (Schonbaum et al, 1995). …”

“…The ovaries were incubated with anti-Yl whole serum (1:100 -1:300, diluted in PBTxT ϩ 2%BSA) or affinity-purified anti-Yl antibodies (1:100) overnight at 4°C. The rat anti-Yl antibodies have been described previously (Schonbaum et al, 1995). The ovaries were washed extensively with four 1.5-h washes with PBTxT and then incubated overnight at 4°C with preabsorbed secondary antibody (fluorescein-conjugated goat anti-rat immunoglobulin, diluted 1:400 -1:500 in PBTxT.…”

Cellular uptake of steroid carrier proteins—Mechanisms and implications

“…Very-low density lipoprotein (VLDL) receptors and yolkless are expressed in oocytes of egg-laying species from insects to birds and mediate the endocytic uptake of yolk (Bujo et al, 1995;Grant and Hirsh, 1999;Schonbaum et al, 1995).…”