Biomedical subjects

K Iatrou

Publications and source records attributed to K Iatrou.

At least 37 records · Page 2Linked to original sources

PubMed1996

The genes encoding the P39 and CG30 proteins of Bombyx mori nuclear polyhedrosis virus.

We have cloned and analysed the transcriptional properties of two closely linked genes, p39 and cg30, of Bombyx mori nuclear polyhedrosis virus (BmNPV). These genes encode a structural polypeptide and a putative transcriptional regulator of the virus, respectively. The cg30 gene is transcribed prior to and after DNA replication from a site located within the ORF of the adjacent p39 gene. Its transcription product, a 1.3 kb mRNA, is polyadenylated at a site containing consensus eukaryotic polyadenylation signals and mapping 87 bp downstream of the translation termination codon for CG30. During the later stages of infection, two additional RNAs, 2.2 and 6.5 kb, are also transcribed through the cg30 gene. The 2.2 kb RNA, representing the mRNA that encodes P39, is initiated from three relatively closely spaced sites located upstream of the P39 ORF. The 6.5 kb RNA is apparently transcribed from the promoter sequences of another gene located further upstream of the p39 gene. The 2.2 and 6.5 kb transcripts have two polyadenylation sites. The first site is the same as the one used to generate the cg30 gene-specific transcripts. The second is located 4 bp downstream of the CG30 translation termination codon. Transient expression assays show that the p39 gene sequences immediately upstream of the CG30 ORF can direct expression of a reporter gene when the latter is co-transfected with the gene encoding the early baculovirus trans-activator IE1. Thus, these sequences behave as a delayed-early baculovirus gene promoter.

Amino Acid Sequence↗

PubMed1995

Developmental regulation of a silkworm gene encoding multiple GATA-type transcription factors by alternative splicing.

Gene BmGATA beta of the silkworm Bombyx mori was previously shown to encode factor BCFI, which regulates the expression of a class of chorion genes expressed during the late stages of choriogenesis. We now show that the expression of the BmGATA beta gene is spatially and temporally regulated by alternative splicing that generates two major (BmGATA beta 1 and BmGATA beta 2) and one minor (BmGATA beta 3) mRNA isoforms of non-identical tissue distribution. The three isoforms differ in the organization of the DNA-binding domains of the corresponding polypeptides. While all three isoforms are expressed in ovarian follicular cells and in testes, only one of them, BmGATA beta 1, is gonad-specific. BmGATA beta 2 is expressed in a variety of other larval and pupal tissues, while BmGATA beta 3 is detected in some pupal but none of the larval tissues. Analysis of RNA isolated from follicular cells of developing ovarian follicles has shown that the onset of ovarian transcription for all three mRNA isoforms occurs during late vitellogenesis, and that the level of accumulated mRNA declines significantly at the onset of choriogenesis. Coincident with the onset of late chorion gene expression, we have observed a significant change in the preference of splice site selection in favour of the one that results in the generation of BmGATA beta 1 mRNA. The transcriptional activation of the BmGATA beta gene in follicular cells during late vitellogenesis correlates with the previously demonstrated initial accumulation of factor BCFI in the cytoplasm of follicular cells and its appearance in follicular cell nuclei only during the late stages of choriogenesis. The relationship between factor BCFI and the different polypeptides encoded by the three BmGATA beta mRNA isoforms is discussed.

Alternative Splicing↗

PubMed1995

The silkmoth homolog of the Drosophila ecdysone receptor (B1 isoform): cloning and analysis of expression during follicular cell differentiation.

To understand the role that 20-hydroxy-ecdysone (20E) plays during ovarian development in Bombyx mori, we have undertaken the cloning of the silkworm ecdysone receptor (EcR) and a study of its expression during follicular cell differentiation. We have cloned a cDNA that contains a complete open reading frame for a 68.1 kDa polypeptide that shares extensive similarities with the B1 isoform of the Drosophila EcR. The presumed silkmoth EcR (BmEcR) is encoded by a single copy gene whose length is in excess of 23 kb. A portion of this gene encompassing seven exons that constitute the cloned BmEcR cDNA was also characterized. Employment of monoclonal antibodies, directed against the DNA binding domain of the Drosophila EcR, in Western blot analyses revealed the presence of a major 70 kDa polypeptide in extracts of follicular cells and other silkmoth tissues. The mRNA and protein encoded by BmEcR are present in constant amounts in follicular cells throughout vitellogenesis but disappear transiently at the onset of choriogenesis and reappear during the later stages of choriogenesis. The down-regulation of BmEcR in follicular cells during oogenesis suggest a complex relationship between 20E, the induction of the program of chorion gene expression in follicular cells during mid-vitellogenesis and the execution of this program at the end of vitellogenesis.

Amino Acid Sequence↗

PubMed1994

Protein phosphorylation and control of chorion gene activation through temporal mobilization of a promoter DNA binding factor from the cytoplasm into the nucleus.

The transcriptional activation of high cysteine chorion genes in the follicular cells of the silkworm Bombyx mori occurs at the end of oogenesis and coincides with the appearance of a chorion promoter DNA binding factor, BCFI, in follicular cell nuclei. Follicular cells of vitellogenic and choriogenic follicles that do not express high cysteine chorion genes contain high levels of a latent form of BCFI in their cytoplasm. The abundance of the cytoplasmic factor, termed cBCFI, is dramatically reduced during late choriogenesis, coincident with the appearance of factor BCFI in the nucleus and the transcriptional activation of high cysteine genes. Mobility shift assays performed with partially proteolyzed nuclear and cytoplasmic extracts of follicular cells, DNA binding assays carried out in the presence of anti-BCFI antibodies, and electrophoretic analyses of the proteins present in the nuclear and cytoplasmic fractions of follicular cells and recognized by the same antibodies suggest that factor cBCFI represents a covalently modified version of BCFI. The DNA-binding sites of BCFI and cBCFI include a core sequence, AGATAA, but, while this sequence is sufficient for specific binding of BCFI, it only constitutes part of the DNA-binding site of cBCFI. Dephosphorylation of cBCFI results in a change of its binding specificity to that of BCFI. The cytoplasmic sequestration of cBCFI appears to be mediated by a phosphorylation-dependent, reversible association of this factor with an ancillary cytoplasmic factor.

Animals↗

PubMed1994

GATA-type zinc finger motif-containing sequences and chorion gene transcription factors of the silkworm Bombyx mori.

To characterize a DNA-binding protein, BCFI, which regulates the expression of silkmoth chorion genes through binding to gene promoter elements identical to those recognized by the GATA family of transcription factors, we have carried out polymerase chain reaction amplifications of Bombyx mori genomic DNA using degenerate primers derived from the conserved DNA binding domain of mammalian GATA factors. Two single copy genes, BmGATA alpha and BmGATA beta, were identified, which encode sequences containing GATA-type zinc finger motifs. The BmGATA beta gene is expressed in follicular and Bm5 tissue culture cells, the two cell types that contain BCFI. No BmGATA alpha gene transcripts were detectable in the tissues that were tested. Upon overexpression in Escherichia coli, a peptide encompassing the BmGATA beta zinc finger motif was able to bind specifically to the BCFI recognition motif of the chorion gene promoters. A polyclonal antibody directed against the zinc finger domain of BmGATA beta was also used in gel retardation assays to confirm that factor BCFI is indeed encoded by the BmGATA beta gene. Conceptual translation of a complete cDNA clone encoding the BmGATA beta protein revealed that this protein has a size similar to that of an immunoreactive protein, presumably BCFI, which is present in follicular cell extracts.

Amino Acid Sequence↗

PubMed1993

Silk moth chorion pseudogenes: hallmarks of genomic evolution by sequence duplication and gene conversion.

The part of the genetic locus of the domesticated silk moth, Bombyx mori, in which high cysteine (Hc) chorion genes of late developmental specificity reside contains regions encompassing gene-like sequences which exhibit properties distinct from those of functional Hc genes. One of these regions has been characterized and shown to contain a chorion pseudogene, psi HcB.15, which shares pronounced similarities with a transcribed chorion pseudogene, psi HcB.12/13, which was characterized previously. Both pseudogenes are homologous to HcB chorion genes but bear multiple single nucleotide substitutions and short segmental mutations (insertions and deletions) which introduce translational frame shifts and termination codons in the coding regions. Structural characteristics unique to the two pseudogenes suggest that psi HcB.15 was generated first from a functional HcB gene and gave rise subsequently to psi HcB 12/13 as a result of a sequence duplication event. The two pseudogenes can be distinguished from each other by the presence of distinct regions of similarity to the consensus sequence of functional HcB genes which appear to have arisen from gene-conversion-mediated correctional events. These findings lend support to the hypothesis that chorion pseudogene sequences represent reservoirs of genetic information that participates in the evolution of the chorion locus rather than relics of inactivated genes passively awaiting extinction.

Animals↗

PubMed1992

Early establishment and autonomous implementation of a developmental program controlling silkmoth chorion gene expression.

To address the question of whether prechoriogenic follicles of the silkmoth Bombyx mori have the capacity to enter choriogenesis in organ culture and define the stage at which choriogenesis becomes established as a follicle-autonomous program, we have cultured immature ovarioles dissected from developing pupae and examined the protein synthetic profiles of follicular cells of individual follicles at the end of the culture period. The protein synthetic profiles of the cultured follicles were also correlated with corresponding profiles of chorion mRNA accumulation. Our results demonstrate that the last 17 (+/- 2) vitellogenic follicles of Day 5 to 7 pupae are capable of initiating choriogenesis in organ culture. The earliest vitellogenic stage to enter choriogenesis in vitro does so after 34 (+/- 4) hr in culture and follicles entering choriogenesis in vitro are capable of proceeding through all choriogenic stages at a speed comparable to that occurring in vivo. Therefore, once the choriogenic program becomes established in follicular cells, it can be implemented autonomously in the absence of extrafollicular factors. Earlier vitellogenic stages lack this capacity, presumably because they require additional hemolymph factors to establish the choriogenic potential. Our results demonstrate that the choriogenic potential of cultured vitellogenic follicles cannot be influenced by addition of 20-hydroxyecdysone to the culture medium.

Animals↗

PubMed1992

A cellular promoter-based expression cassette for generating recombinant baculoviruses directing rapid expression of passenger genes in infected insects.

We have developed an expression cassette which allows the generation of recombinant baculoviruses that can express passenger genes under the control of a constitutive cellular promoter derived from the cytoplasmic actin gene of the silkmoth Bombyx mori. Silkmoth tissue culture cells which were infected with a recombinant B. mori nuclear polyhedrosis virus (BmNPV) containing the gene-encoding chloramphenicol acetyltransferase (CAT) under the control of this expression cassette expressed significant CAT activity beginning 5 hr postinfection (p.i.). Cells infected with a recombinant BmNPV containing the cat gene under the control of the polyhedrin gene promoter did not express CAT activity until 20 hr p.i. Silkworm larvae were also infected with the two recombinant viruses by hemocelic injections and all larval tissues examined were found to express the cat gene. While significant actin-cassette-driven CAT expression in vivo was first seen at 24 hr p.i., expression from the polyhedrin promoter was not seen until 48 hr p.i. By 60 hr p.i., tissues of larvae infected with the recombinant virus expressing cat under polyhedrin promoter control were found to exhibit sixfold higher CAT activity than those infected with recombinant virus expressing the cat gene under the control of the actin promoter. The 24-hr temporal advantage in expression of a passenger gene in infected larvae indicates that the actin-promoter-based expression cassette or other analogous cellular promoter-based cassettes could be used for generating recombinant baculovirus insecticides which could incapacitate pest insects more quickly than viruses employing the polyhedrin or other late viral promoters for expressing insect-incapacitating proteins.

Actins↗

PubMed1991

Developmental regulation of covalent modification of double-stranded RNA during silkmoth oogenesis.

Follicular cells of the silkmoth Bombyx mori contain an enzymatic activity that modifies RNA duplexes in vitro. The modifying activity converts adenosine residues into inosine in duplex but not single-stranded RNA and mediates the partial unwinding of the complement strands. Because of the modification, the RNA loses its ability to form perfect duplexes with its complement upon reannealing in vitro. The modifying enzyme is localized in the cytoplasm of follicular cells and its activity is modulated in a developmentally regulated manner. In contrast, follicular nuclei contain an activity that inhibits the modification and unwinding of duplex RNA. The modifying activity is also present in the cytoplasm of unfertilized oocytes and its accumulation during oogenesis parallels that of the follicular cells. Examination of an established silkmoth cell line of ovarian origin revealed that, in contrast to the situation with follicular cells, the modifying activity has an exclusive nuclear localization. The cytoplasmic fraction of these cells is not only devoid of modifying activity but, as is the case with the nuclear fraction of follicular cells, contains an activity that inhibits duplex RNA modification and unwinding. We conclude that the modification promoting and inhibiting activities are not restricted to a single cell type and that their compartmentalization is developmentally regulated.

Adenosine↗

PubMed1991

Synergistic interactions of silkmoth chorion promoter-binding factors.

Two DNA-binding proteins, BCFI and BCFII, that interact with defined promoter sequences of silkmoth chorion genes of late developmental specificity appear in the nuclei of follicular cells at a time that coincides with the transcriptional activation of the corresponding genes. BCFI prebinding is shown to be indispensable for stable binding of BCFII to its cognate sequence. BCFI and BCFII synergism requires a relatively stringent stereospecific alignment and is a prerequisite for the assembly of higher-order protein-promoter DNA complexes containing additional factors, which are neither gene (stage) nor class (chorion) specific. Binding of BCFI to its site correlates with the induction of DNA structural perturbations that may facilitate assembly of additional factors on the promoter. The BCFI-binding domain contains a core hexanucleotide sequence, AGATAA, which represents the major binding determinant of the erythroid-specific transcription factor GATA-1 of higher vertebrates. This sequence is shown to be necessary and sufficient for binding of BCFI, as it is for a factor that is present in induced K562 human erythroleukemic cells, presumably GATA-1. Comparative analyses of mobility shift patterns obtained with partially proteolyzed preparations of these two unrelated factors were used to confirm that a BCFI-like chorion promoter-binding protein, which is present in the nuclei of an established silkmoth cell line derived from ovarian tissue, is in fact BCFI. The transcriptional repression of endogenous chorion genes in this cell line coupled with the documented absence of factor BCFII suggests that the synergistic interactions between these two factors constitute a minimum requirement for late chorion gene expression.

Animals↗

PubMed1990

Silkmoth chorion antisense RNA. Structural characterization, developmental regulation and evolutionary conservation.

Choriogenic follicular cells of the silkmoth Bombyx mori contain significant quantities of antisense RNA transcribed from chorion genes. Antisense RNA derived from a chorion gene with a high content of cysteine, HcB.12, was characterized in detail. The antisense transcripts are initiated downstream from the 3' end of HcB.12 mRNA and extend over 75% of the length of the gene, comprising its entire second exon and part of its intervening sequence. The antisense RNA is devoid of any significant open reading frames and is not polyadenylated. These features, combined with the presence of specific sequence motifs within its transcribed and upstream region, suggest that antisense RNA may be transcribed by RNA polymerase III. Chorion antisense RNA is detectable only in choriogenic follicular cells and appears to be co-ordinately regulated with chorion mRNA. Its cytoplasmic accumulation during choriogenesis parallels that of the corresponding mRNA. Although chorion mRNA is at least five times more abundant than antisense RNA, the latter is present as a single-stranded entity in follicular cytoplasm but can form perfect duplexes with its mRNA complement upon annealing in vitro. The possible involvement of antisense RNA transcription in the pathway that controls the programmed expression of chorion genes at the level of transcription initiation or post-transcriptional processing is discussed.

Animals↗

PubMed1990

Tissue-specific expression of silkmoth chorion genes in vivo using Bombyx mori nuclear polyhedrosis virus as a transducing vector.

A pair of silkmoth chorion chromosomal genes, HcA.12-HcB.12, was inserted into a baculovirus transfer vector, pBmp2, derived from the nuclear polyhedrosis virus of Bombyx mori. This vector, which permits the insertion of foreign genetic material in the vicinity of a mutationally inactivated polyhedrin gene, was used to acquire the corresponding recombinant virus. Injection of mutant silkmoth pupae that lack all Hc chorion genes with the recombinant virus resulted in the infection of all internal organs including follicular tissue. Analysis of RNA from infected tissues has demonstrated that the two chorion genes present in the viral genome are correctly transcribed under the control of their own promoter in follicular cells, the tissue in which chorion genes are normally expressed. The chorion primary transcripts are also correctly processed in the infected follicular cells and yield mature mRNAs indistinguishable from authentic chorion mRNAs present in wild-type follicles. These results demonstrate that recombinant nuclear polyhedrosis viruses can be used as transducing vectors for introducing genetic material of host origin into the cells of the organism and that the transduced genes are transiently expressed in a tissue-specific manner under the control of their resident regulatory sequences. Thus we show the in vivo expression of cloned genes under cellular promoter control in an insect other than Drosophila melanogaster. The approach should be applicable to all insect systems that are subject to nuclear polyhedrosis virus infection.

Animals↗

PubMed1989

GrB deletion of the chorion locus of the silkmoth Bombyx mori. Localization of the left breakpoint and isolation of the deletion junction.

In the silkmoth Bombyx mori, choriogenesis occurs through the developmentally controlled deposition of several related classes of chorion proteins onto the oocyte by surrounding follicular cells. In the GrB mutant strain, a distinctive family of proteins (Hc) normally expressed late in choriogenesis, as well as several proteins of middle development specificity, are missing due to the deletion of the corresponding genes from the chorion locus. In addition, a smaller set of proteins normally confined to mid-choriogenesis is found to be prolonged in expression in homozygote mutant but not heterozygote individuals. To elucidate the molecular organization of the chorion locus in the GrB genotype, we scanned a part of the wild-type locus represented by a chromosomal walk of 270,000 bases through library screening and genomic DNA hybridizations using a series of unique probes. A chromosomal clone, GrB4, whose sequences showed the expected characteristics of the deletion junction, was isolated from a partial EcoRI library of mutant genomic DNA. Through comparative hybridizations, mapping and sequencing, the precise location of one of the deletion breakpoints was identified on one of the clones mapping in the characterized part of the wild-type locus. Attempts to locate the other breakpoint in wild-type DNA and to extend the structural characterization past the deletion junction through chromosomal walking were unsuccessful, due to the apparent absence of these sequences from libraries of wild-type and mutant genomic DNA, respectively. Hybridizations of the deletion region on clone GrB4 to cDNA derived from follicular RNA indicate that no gene sequences are directly interrupted by the deletion, and reveal the presence of a gene sequence of unknown function 1000 to 5000 bases to the right of deletion junction.

Animals↗

PubMed1989

Bombyx mori nuclear polyhedrosis virus-based vectors for expressing passenger genes in silkmoth cells under viral or cellular promoter control.

The polyhedrin gene of the nuclear polyhedrosis virus of the silkmoth Bombyx mori (BmNPV) has been subjected to deletion mutagenesis. A number of clones containing partially deleted polyhedrin genes were characterized and four clones containing limited deletions of the 5'-untranslated or 5'-flanking sequences of the gene were further analyzed with respect to polyhedrin promoter activity. The functional characterization of the deletion mutants was achieved through the insertion of a chloramphenicol acetyl transferase (CAT) gene (cat) into each deletion junction. The resultant cat constructs were introduced into the genome of BmNPV through homologous recombination and the effect of each deletion on the activity of the polyhedrin promoter was evaluated by measurements of CAT enzymatic activity in extracts of tissue culture cells infected with the corresponding recombinant BmNPVs as well as by primer extension assays. Removal of the entire leader region and eleven adjacent residues of the 5'-flanking sequences of the polyhedrin gene results in a dramatic decrease in promoter activity, which, however, remains detectable through CAT activity measurements. Elimination of an additional 30 nucleotides (nt) of the upstream sequences results in the complete inactivation of the polyhedrin promoter. The functional characterization of a deletion mutant lacking 41 nt of the 5'-flanking sequences has demonstrated that no functions necessary for viral infectivity, replication or assembly are disrupted by this deletion, since the corresponding recombinant viruses propagate in the cells with the same kinetics and to the same extent as wild-type BmNPV. As a result of the deletion mutagenesis, two classes of transfer vectors have become available. The first class can be used for introducing into the viral genome foreign nucleotide sequences under polyhedrin promoter control, while the second one can be used for obtaining recombinant viruses harboring foreign genetic material in an environment which is devoid of polyhedrin promoter activity.

Amino Acid Sequence↗

PubMed1989

Recombinant baculoviruses as vectors for identifying proteins encoded by intron-containing members of complex multigene families.

Using a transfer vector derived from Bombyx mori nuclear polyhedrosis virus (BmNPV), we have constructed recombinant baculoviruses that contain complete silk moth chorion chromosomal genes encoding high-cysteine proteins under the control of the polyhedrin promoter. Silk moth tissue culture cells infected with these recombinant viruses were found to contain abundant RNA sequences of sizes similar to those of the authentic chorion mRNAs. Chorion transcripts present in infected cells were initiated almost exclusively at the cap site of the polyhedrin start site. Primer extension and RNase protection experiments revealed that a considerable proportion of the resultant transcripts were spliced at the same sites as those utilized in follicular cells for the production of functional chorion mRNA. Electrophoretic analysis and immunoprecipitation of the proteins of host cells infected with the recombinant viruses revealed the presence of the corresponding chorion proteins. We conclude that baculovirus vectors can be used for expressing efficiently not only cDNAs or simple genes devoid of intervening sequences but also intron-containing chromosomal genes. Thus, recombinant baculoviruses offer a powerful alternative to hybrid-selected translation, particularly when the identification of proteins encoded by members of complex multigene families is required.

Amino Acid Sequence↗

PubMed1988

Identification of a transcriptionally active pseudogene in the chorion locus of the silkmoth Bombyx mori. Regional sequence conservation and biological function.

We have determined the primary structure of a 3500 base-pair part of the silkmoth chorion locus mapping in a region containing genes of late developmental specificity. This part of the locus was found to harbour a pseudogene related to one of the families of chorion genes encoding high cysteine proteins, HcB. The pseudogene exhibits an overall sequence identity of 84% to the consensus coding region of HcB chorion genes. A 95% identity was observed over a length of 190 base-pairs of its immediate 5' upstream sequences and the corresponding part of the consensus 5'-intergenic sequences of Hc gene pairs, normally encompassing 270 base-pairs. Thus, the pseudogene has retained part of the promoter region that includes sequence elements whose presence is thought to be necessary for transcriptional competence of HcB genes. The pseudogene is also characterized by the elimination of part of its first exon containing most of the 5' untranslated region, the ATG translation initiation codon and part of the signal peptide sequences. Its intron is longer than that of other HcB genes due to the insertion of a copy of a repetitive element that appears to be transcribed by RNA polymerase III. A previously characterized chorion cDNA clone, m2282, representing a rare mRNA sequence of late developmental specificity, was found to be identical to the pseudogene over its entirety spanning 65% of the pseudogene's second exon. Hybridizations of clones spanning a 260,000 base-pair domain of the chorion locus of Bombyx mori and of total genomic DNA to a subfragment of the cDNA clone containing relatively unique sequences, coupled to primer extension experiments, have demonstrated that m2282 mRNA originated from the pseudogene and that the pseudogene transcripts are initiated at the chorion cap site consensus sequence. We conclude that the 5'-flanking sequences retained by the pseudogene encompass elements necessary and adequate for correct transcriptional activation, but may not include those required for quantitative expression of the promoter. Possible reasons for the observed lack of random drift in the 5'-upstream sequences of the pseudogene and the maintenance of a functional promoter in a non-functional gene are discussed on the basis of the observation that elements resembling scaffold attachment sites are present in these sequences.

Amino Acid Sequence↗

PubMed1987

Transcriptional behavior of silkmoth chorion genes in vivo and in injected Xenopus laevis oocytes.

A cloned fragment of Bombyx mori genomic DNA containing a pair of chorion genes was injected into Xenopus laevis oocytes. The RNA transcribed from the injected template was examined by primer extension analysis for the presence of transcripts initiated correctly from the two chorion genes. Using as primers a number of fragments derived from various parts of the injected DNA, we have found that transcription initiation around the cap sites of the chorion genes was considerably more frequent than that at other, seemingly random sites of the template. Injection of different quantities of template into the oocytes revealed that at high DNA concentrations, most transcripts were randomly initiated. However, when reduced amounts of DNA were injected into the oocytes, the extent of specific transcription increased dramatically while the proportion of nonspecific transcripts correspondingly decreased. The relative rates of specific initiation from the two chorion genes in injected oocytes resembled closely those observed in choriogenic follicles. Finally, the primer extension analysis revealed the presence of chorion antisense transcripts not only in injected oocytes but also in total RNA derived from follicular cells. The possible origin of these transcripts is discussed.

Animals↗

PubMed1985

Polyhedrin gene of Bombyx mori nuclear polyhedrosis virus.

A portion of the genome of the nuclear polyhedrosis virus of Bombyx mori has been cloned. This part of the viral genome contains the gene encoding the viral occlusion body protein, polyhedrin. The polyhedrin gene has been sequenced in its entirety together with some of its 5' and 3' flanking sequences. The primary structure of polyhedrin predicted from the nucleotide sequence of the gene was found to be somewhat different from the one reported previously for the authentic protein (E. A. Kozlov, T. L. Levitina, N. M. Gusak, and S. B. Serebryani, Bioorg. Khim., 7:1008-1015, 1981; S. B. Serebryani, T. L. Levitina, M. L. Kautsman, Y. L. Radavski, N. M. Gusak, M. N. Ovander, N. V. Sucharenko, and E. A. Kozlov, J. Invertebr. Pathol., 30:442-443, 1977). Comparison of the primary structures of the polyhedrin of the nuclear polyhedrosis virus of B. mori with that of Autographa californica suggests that considerable selective pressure has been exercised at the protein level during evolution. Nucleotide sequence comparisons of the two structural genes reveal that the coding sequences have diverged significantly through the accumulation of silent and replacement substitutions. In contrast, a remarkable degree of sequence conservation was found to exist in the domains corresponding to the 5' and 3' noncoding regions of the polyhedrin mRNAs.

Amino Acid Sequence↗