Molecular characterization and gene expression in the eye of the apolipophorin II/I precursor fromLocusta migratoria

Abstract

The transport of lipids via the circulatory system of animals constitutes a vital function that uses highly specialized lipoprotein complexes. In insects, a single lipoprotein, lipophorin, serves as a reusable shuttle for the transport of lipids between tissues. We have found that the two nonexchangeable apolipoproteins of lipophorin arise from a common precursor protein, apolipophorin II/I (apoLp-II/I). To examine the mechanisms of transport of lipids and liposoluble substances inside the central nervous system, this report provides the molecular cloning of a cDNA encoding the locust apoLp-II/I. We have recently shown that this precursor protein belongs to a superfamily of large lipid transfer proteins (Babin et al. [1999] J. Mol. Evol. 49:150-160). We determined that, in addition to its expression in the fat body, the locust apoLp-II/I is also expressed in the brain. Part of the signal resulted from fat body tissue associated with the brain; however, apoLp-II/I was strongly expressed and the corresponding protein detected, in pigmented glial cells of the lamina underlying the locust retina and in cells or cellular processes interspersed in the basement membrane. The latter finding strongly suggests an implication of apolipophorins in the transport of retinoids and/or fatty acids to the insect retina.

Biosynthesis and secretion of insect lipoprotein: involvement of furin in cleavage of the apoB homolog, apolipophorin-II/I

“…It is well established that insect lipoproteins contain two apolipoproteins, apoLp-I and apoLp-II, that are derived from their common precursor protein apoLp-II/I (6,(8)(9)(10)(11). In the present study, we identified and characterized the cleavage site in L. migratoria apoLp-II/I using an insect recombinant expression system of Sf9 cells and demonstrated that its cleavage likely involves an insect furin.…”

“…I), which is similar to that reported for wild-type locust apoLp-II (6). Moreover, the similar migration behavior of recombinant and wild-type apoLp-II indicates that the cleavage of apoLp-II/I-38 proceeds identical to that in fat body (6,10). Expression of a construct expressing the N-terminal 33% of apoLp-II/I in Sf9 cells, and apoLp-II/I-38 in Drosophila melanogaster S2 cells, also resulted in the secretion of the two expected apoLp-II/I cleavage products (data not shown).…”

“…Standard cloning and sequencing procedures (10,14) were used to obtain an expression construct for L. migratoria apoLp-II/I. The apoLp-II/I-38 truncation variant was constructed by cloning the large Eco RI-Xho I cDNA fragment from pBK-CMV cDNA clone B20 (10) into pGEM7-Zf ϩ (Promega).…”

“…1A ). Excluding the 21 residue signal peptide (10), this apoLp-II/I truncation variant corresponds to 38% of the total precursor protein and is referred to as apoLp-II/I-38.…”

Lipophorin receptor-mediated lipoprotein endocytosis in insect fat body cells

“…HDLp comprises diacylglycerol and phospholipids as major lipid classes. The protein matrix consists of two nonexchangeable apolipoproteins, apolipophorin I (apoLp-I) and apoLp-II, which are derived from a common precursor protein through posttranslational cleavage (3,4). Sequence and domain structure analyses indicate that this precursor protein is homologous to mammalian apolipoprotein B-100 (apoB-100), the nonexchangeable protein component of VLDL and its resulting LDL, and that both proteins have emerged from an ancestral gene (5)(6)(7).…”