Biomedical subjects

K Mita

Publications and source records attributed to K Mita.

At least 19 recordsLinked to original sources

PubMed2006

Isolation and characterization of sex chromosome rearrangements generating male muscle dystrophy and female abnormal oogenesis in the silkworm, Bombyx mori.

In deletion-mapping of W-specific RAPD (W-RAPD) markers and putative female determinant gene (Fem), we used X-ray irradiation to break the translocation-carrying W chromosome (W( Ze )). We succeeded in obtaining a fragment of the W( Ze ) chromosome designated as Ze (W), having 3 of 12 W-RAPD markers (W-Bonsai, W-Yukemuri-S, W-Yukemuri-L). Inheritance of the Ze (W) fragment by males indicates that it does not include the Fem gene. On the basis of these results, we determined the relative positions of W-Yukemuri-S and W-Yukemuri-L, and we narrowed down the region where Fem gene is located. In addition to the Ze (W) fragment, the Z chromosome was also broken into a large fragment (Z(1)) having the +( sch ) (1-21.5) and a small fragment (Z(2)) having the +( od ) (1-49.6). Moreover, a new chromosomal fragment (Ze (W)Z(2)) was generated by a fusion event between the Ze (W) and the Z(2) fragments. We analyzed the genetic behavior of the Z(1) fragment and the Ze (W)Z(2) fragment during male (Z/Z(1) Ze (W)Z(2)) and female (Z(1) Ze (W)Z(2)/W) meiosis using phenotypic markers. It was observed that the Z(1) fragment and the Z or the W chromosomes separate without fail. On the other hand, non-disjunction between the Ze (W)Z(2) fragment and the Z chromosome and also between the Ze (W)Z(2) fragment and the W chromosome occurred. Furthermore, the females (2A: Z/Ze (W)Z(2)/W) and males (2A: Z/Z(1)) resulting from non-disjunction between the Ze (W)Z(2) fragment and the W chromosome had phenotypic defects: namely, females exhibited abnormal oogenesis and males were flapless due to abnormal indirect flight muscle structure. These results suggest that Z(2) region of the Z chromosome contains dose-sensitive gene(s), which are involved in oogenesis and indirect flight muscle development.

Animals↗

PubMed2006

The female-killing chromosome of the silkworm, Bombyx mori, was generated by translocation between the Z and W chromosomes.

Bombyx mori is a female-heterogametic organism (female, ZW; male, ZZ) that appears to have a putative feminizing gene (Fem) on the W chromosome. The paternally transmitted mutant W chromosome, Df(p ( Sa ) + ( p )W + ( od ))Fem, derived from the translocation-carrying W chromosome (p ( Sa ) + ( p )W + ( od )), is inert as femaleness determinant. Moreover, this Df(p ( Sa ) + ( p )W + ( od ))Fem chromosome has been thought to have a female-killing factor because no female larvae having the Df(p ( Sa ) + ( p )W + ( od ))Fem chromosome are produced. Initially, to investigate whether the Df(p ( Sa ) + ( p )W + ( od ))Fem chromosome contains any region of the W chromosome or not, we analyzed the presence or absence of 12 W-specific RAPD markers. The Df(p ( Sa ) + ( p )W + ( od ))Fem chromosome contained 3 of 12 W-specific RAPD markers. These results strongly indicate that the Df(p ( Sa ) + ( p )W + ( od ))Fem chromosome contains the region of the W chromosome. Moreover, by using phenotypic and molecular markers, we confirmed that the Df(p ( Sa ) + ( p )W + ( od ))Fem chromosome is connected with a partially deleted Z chromosome and that this fused chromosome behaves as a Z chromosome during male meiosis. Furthermore, we demonstrated that the ZZW-type triploid female having the Df(p ( Sa ) + ( p )W + ( od ))Fem chromosome is viable. Therefore, we concluded that the Df(p ( Sa ) + ( p )W + ( od ))Fem chromosome does not have a female-killing factor but that partial deletion of the Z chromosome causes the death of the ZW-type diploid female having the Df(p ( Sa ) + ( p )W + ( od ))Fem chromosome. Additionally, our results of detailed genetic analyses strongly indicate that the female-killing chromosome composed of the Df(p ( Sa ) + ( p )W + ( od ))Fem chromosome and deleted Z chromosome was generated by translocation between the Z chromosome and the translocation-carrying W chromosome, p ( Sa ) + ( p )W + ( od ).

Animals↗

PubMed2006

Cloning and expression analysis of takeout/JHBP family genes of silkworm, Bombyx mori.

A Bombyx EST cDNA database was searched using the Drosophila takeout gene and nine cDNAs were obtained. The homology search suggested that these genes are widespread in insects and organize a large gene family, and that they have hydrophobic ligands. A phylogenetic tree indicated that the genes are first divided into two large groups, juvenile hormone binding protein and other protein genes, and the latter group diversified within a short time at an early stage. The expression study of five Bombyx genes indicated that they are expressed in various tissues and are regulated by development and feeding conditions. The Bombyx genes might have roles related to the regulation of metabolism, growth or development related to nutritional conditions.

Amino Acid Sequence↗

PubMed2006

Retroperitoneoscopic radical nephrectomy in obese patients: outcomes and considerations.

OBJECTIVE: To determine whether obesity is associated with surgical outcome in Japanese patients undergoing retroperitoneoscopic radical nephrectomy (RRN). PATIENTS AND METHODS: Between November 1999 and March 2005, we performed 98 RRN procedures for patients with renal cell carcinoma. Patients with a body mass index (BMI) of 25.0 or more were defined as obese (group A, n=33) and those with a BMI of <25.0 were defined as non-obese (group B, n=65), in accordance with the criteria of the Japan Society for the Study of Obesity. Patient background, degree of surgical invasiveness, and period of convalescence were compared between groups A and B. RESULTS: No statistically significant differences were observed between the groups in terms of age, gender, tumor laterality, tumor size, and time until resumption of oral intake and ambulation. However group A had a significantly longer insufflation time (172.1 vs. 137.4 min), greater blood loss (195.3 vs. 48.4 ml) and higher renal specimen weight (440.0 vs. 306.0 g) than group B. CONCLUSION: Obesity is not a factor that affects patient eligibility for RRN, but is a risk factor for longer insufflation time and greater blood loss.

Carcinoma, Renal Cell↗

PubMed2006

Stat5 expression predicts response to endocrine therapy and improves survival in estrogen receptor-positive breast cancer.

Constitutively activated signal transducers and activators of transcription (Stats), in particular Stat3 and Stat5, have been demonstrated to directly contribute to oncogenesis by stimulating cell proliferation and preventing apoptosis in various cancers. Stat3 is essential in mammary gland epithelial cell apoptosis and involution, whereas Stat5 is well established as a key factor in mammary epithelial cell growth and differentiation. Crosstalk between Stats and estrogen receptor (ER) has been demonstrated by several laboratories and we have focused on the role of Stat5 in ER-positive breast cancer. Using immunohistochemical techniques, we examined the expression of Stat3 and Stat5 in 517 human breast cancer tissues and analyzed their significance for prognosis and prediction of response to endocrine therapy. Stat5 expression was significantly correlated with histological grade (P<0.0001), ER (P=0.02), and progesterone receptor (P=0.026) expression. There was no difference between Stat3 expression and clinicopathological factors. In 346 patients with ER-positive breast cancer, patients with Stat5 positive tumors had significantly increased overall survival (P=0.0009) in multivariate analysis. There were 70 patients who received endocrine therapy as first-line treatment for metastatic breast cancer at relapse. The patients whose primary breast tumors were Stat5 positive, had significantly better response to endocrine therapy (P=0.04), and longer survival after relapse (P=0.0003), than those whose tumors were Stat5 negative. The present study demonstrates for the first time that Stat5 is a predictive factor for endocrine therapy response and a strong prognostic molecular marker in ER-positive breast cancer. Our data suggest that the expression of Stat5 is helpful in selecting patients who may benefit from endocrine therapy.

Adult↗

PubMed2005

Mechanomyographic determination of post-activation potentiation in myopathies.

OBJECTIVE: There is a need to provide an index of muscle contractility in evaluating myopathies especially in the clinical setting. This study was conducted to investigate if the mechanomyogram post-activation potentiation (MMG-PAP) can be used as an index of muscle contractile force potentiation (force-PAP), if it differs between normal and myopathic muscles, and if it can reflect abnormalities in muscle fiber anatomy. METHODS: The correlation between MMG-PAP and force-PAP was evaluated in 12 normal subjects after maximum voluntary contraction (MVC) of the biceps brachii muscle. The same method was then applied to study MMG-PAP in 16 patients with myopathies, 16 disease and 25 normal controls. Mean fiber diameters and the proportions of type 1 and 2 fibers in biopsied biceps brachii muscle were determined and compared with MMG-PAP values. RESULTS: There was a significant positive correlation between force-PAP (197 +/- 148%) and MMG-PAP (135 +/- 68%) immediately after MVC (P < 0.05). The mean MMG-PAP in myopathies (66 +/- 53%) was significantly lower than those of the disease (128 +/- 34%; P < 0.005) and normal controls (120 +/- 56%; P < 0.005). Patients with non-dystrophic myopathies, including those with myositis, had significantly lower MMG-PAP values (38 +/- 20%; P < 0.005) than those with muscular dystrophy (148 +/- 23%). MMG-PAP did not clearly correlate with either type 2 fiber atrophy or type 2 fiber disproportion based on muscle biopsy analysis of myopathic patients. CONCLUSIONS: This study shows that MMG-PAP can be used as an index of muscle contractility and that it is significantly lower in non-dystrophic myopathies compared to normal subjects. MMG-PAP does not seem to reflect abnormal muscle fiber anatomy. SIGNIFICANCE: MMG-PAP may become a valuable non-invasive tool in augmenting routine clinical electrophysiologic studies especially in evaluating muscle contractility in myopathies.

Adult↗

PubMed2005

SilkSatDb: a microsatellite database of the silkworm, Bombyx mori.

The SilkSatDb (silkmoth microsatellite database) (http://www.cdfd.org.in/silksatdb) is a relational database of microsatellites extracted from the available expressed sequence tags and whole genome shotgun sequences of the silkmoth, Bombyx mori. The database has been rendered with a simple and robust web-based search facility, developed using PHP. The SilkSatDb also stores information on primers developed and validated in the laboratory. Users can retrieve information on the microsatellite and the protocols used, along with informative figures and polymorphism status of those microsatellites. In addition, the interface is coupled with Autoprimer, a primer-designing program, using which users can design primers for the loci of interest.

Animals↗

PubMed2005

Partial deletions of the W chromosome due to reciprocal translocation in the silkworm Bombyx mori.

In the silkworm, Bombyx mori (female, ZW; male, ZZ), femaleness is determined by the presence of a single W chromosome, irrespective of the number of autosomes or Z chromosomes. The W chromosome is devoid of functional genes, except the putative female-determining gene (Fem). However, there are strains in which chromosomal fragments containing autosomal markers have been translocated on to W. In this study, we analysed the W chromosomal regions of the Zebra-W strain (T(W;3)Ze chromosome) and the Black-egg-W strain (T(W;10)+(w-2) chromosome) at the molecular level. Initially, we undertook a project to identify W-specific RAPD markers, in addition to the three already established W-specific RAPD markers (W-Kabuki, W-Samurai and W-Kamikaze). Following the screening of 3648 arbitrary 10-mer primers, we obtained nine W-specific RAPD marker sequences (W-Bonsai, W-Mikan, W-Musashi, W-Rikishi, W-Sakura, W-Sasuke, W-Yukemuri-L, W-Yukemuri-S and BMC1-Kabuki), almost all of which contained the border regions of retrotransposons, namely portions of nested retrotransposons. We confirmed the presence of eleven out of twelve W-specific RAPD markers in the normal W chromosomes of twenty-five silkworm strains maintained in Japan. These results indicate that the W chromosomes of the strains in Japan are almost identical in type. The Zebra-W strain (T(W;3)Ze chromosome) lacked the W-Samurai and W-Mikan RAPD markers and the Black-egg-W strain (T(W;10)+(w-2) chromosome) lacked the W-Mikan RAPD marker. These results strongly indicate that the regions containing the W-Samurai and W-Mikan RAPD markers or the W-Mikan RAPD marker were deleted in the T(W;3)Ze and T(W;10)+(w-2) chromosomes, respectively, due to reciprocal translocation between the W chromosome and the autosome. This deletion apparently does not affect the expression of Fem; therefore, this deleted region of the W chromosome does not contain the putative Fem gene.

Animals↗

PubMed2005

Retrotransposable elements on the W chromosome of the silkworm, Bombyx mori.

The sex chromosomes of the silkworm, Bombyxmori, are designated ZW(XY) for females and ZZ(XX) for males. The W chromosome of B. mori does not recombine with the Z chromosome and autosomes and no genes for morphological characters have been mapped to the W chromosome as yet. Furthermore, femaleness is determined by the presence of a single W chromosome, regardless of the number of autosomes or Z chromosomes. To understand these interesting features of the W chromosome, it is necessary to analyze the W chromosome at the molecular biology level. Initially to isolate DNA sequences specific for the W chromosome as randomly amplified polymorphic DNA (RAPD) markers, we compared the genomic DNAs between males and females by PCR with arbitrary 10-mer primers. To the present, we have identified 12 W-specific RAPD markers, and with the exception of one RAPD marker, all of the deduced amino acid sequences of these W-specific RAPD markers show similarity to previously reported amino acid sequences of retrotransposable elements from various organisms. After constructing a genomic DNA lambda phage library of B. mori we obtained two lambda phage clones, one containing the W-Kabuki RAPD sequence and one containing the W-Samurai RAPD sequence and found that these DNA sequences comprised nested structures of many retrotransposable elements. To further analyze the W chromosome, we obtained 14 W-specific bacterial artificial chromosome (BAC) clones from three BAC libraries and subjected these clones to shotgun sequencing. The resulting assembly of sequences did not produce a single contiguous sequence due to the presence of many retrotransposable elements. Therefore, we coupled PCR with shotgun sequencing. Through these analyses, we found that many long terminal repeat (LTR) and non-LTR retrotransposons, retroposons, DNA transposons and their derivatives, have accumulated on the W chromosome as strata. These results strongly indicate that retrotransposable elements are the main structural component of the W chromosome.

Animals↗

PubMed2005

Interferon-beta induction/interferon-alpha2b plus ribavirin therapy in patients with chronic hepatitis C.

Treatment of chronic hepatitis C virus (HCV) infection with interferon (IFN) and ribavirin improves the rate of eradication of the virus by less than 20% in patients with genotype 1b and a high viral load. In this study we assessed whether IFN-beta induction/IFN-alpha2b plus ribavirin enhances the efficacy of the therapy in patients with chronic hepatitis C. The efficacy of IFN-beta induction/IFN-alpha2b plus ribavirin therapy (group A, n=7) was compared with that of IFN-alpha2b plus ribavirin (group B, n=7) in 14 patients with high levels of HCV-RNA (> 100 K/U/ml). No significant differences were observed in the clearance of HCV-RNA between the two groups (A and B, respectively) 2 weeks after the start of the treatment (0% and 14.3%), at the end of the treatment (71.4% and 100%) and 6 months after the end of the treatment (28.6% and 14.3%). Recovery was complete in 28.6% and 14.3%, transient in 42.9% and 85.7% and absent in 28.6% and 0% in groups A and B, respectively. Early log changes in the viral load from the baseline after 2 weeks of treatment were 2.41 +/- 0.91 and 2.77 +/- 0.20 in groups A and B, respectively, with no significant difference between the two groups. In the present study, we were not able to demonstrate that IFN-beta induction/IFN-alpha2b plus ribavirin therapy was superior to IFN-alpha2b plus ribavirin therapy in patients with genotype 1b and high viral loads.

Adult↗

PubMed2004

Survey and analysis of microsatellites in the silkworm, Bombyx mori: frequency, distribution, mutations, marker potential and their conservation in heterologous species.

We studied microsatellite frequency and distribution in 21.76-Mb random genomic sequences, 0.67-Mb BAC sequences from the Z chromosome, and 6.3-Mb EST sequences of Bombyx mori. We mined microsatellites of >/=15 bases of mononucleotide repeats and >/=5 repeat units of other classes of repeats. We estimated that microsatellites account for 0.31% of the genome of B. mori. Microsatellite tracts of A, AT, and ATT were the most abundant whereas their number drastically decreased as the length of the repeat motif increased. In general, tri- and hexanucleotide repeats were overrepresented in the transcribed sequences except TAA, GTA, and TGA, which were in excess in genomic sequences. The Z chromosome sequences contained shorter repeat types than the rest of the chromosomes in addition to a higher abundance of AT-rich repeats. Our results showed that base composition of the flanking sequence has an influence on the origin and evolution of microsatellites. Transitions/transversions were high in microsatellites of ESTs, whereas the genomic sequence had an equal number of substitutions and indels. The average heterozygosity value for 23 polymorphic microsatellite loci surveyed in 13 diverse silkmoth strains having 2-14 alleles was 0.54. Only 36 (18.2%) of 198 microsatellite loci were polymorphic between the two divergent silkworm populations and 10 (5%) loci revealed null alleles. The microsatellite map generated using these polymorphic markers resulted in 8 linkage groups. B. mori microsatellite loci were the most conserved in its immediate ancestor, B. mandarina, followed by the wild saturniid silkmoth, Antheraea assama.

Animals↗

PubMed2004

Hormonal control of a metamorphosis-specific transcriptional factor Broad-Complex in silkworm.

Insect metamorphosis is induced by the steroid 20-hydroxyecdysone (20E) in the absence of sesquiterpenoid juvenile hormone (JH). In Drosophila melanogaster, the Broad-Complex (BR-C) transcriptional factor plays critical roles during metamorphosis. We isolated cDNAs encoding BR-C in the silkworm Bombyx mori and examined their mRNA expression. cDNAs for three BR-C isoforms with zinc finger pairs (Z1, Z2 and Z4) and four isoforms lacking them were cloned. Their mRNAs were expressed in multiple tissues at the larval-pupal metamorphosis. In the anterior silk gland, BR-C mRNAs were expressed at the end of the last larval instar but not expressed during the penultimate instar. 20E administration induced BR-C mRNA expression and JH suppressed 20E-induced BR-C expression in this tissue both in vivo and in vitro. Thus, BR-C mRNAs are inducible by 20E only in the absence of JH, a finding that explains their metamorphosis-specific expression.

Animals↗

PubMed2004

Accumulation of spermidine/spermine N1-acetyltransferase and alternatively spliced mRNAs as a delayed response of HeLa S3 cells following X-ray irradiation.

PURPOSE: A key enzyme of polyamine catabolism, spermidine/spermine N(1)-acetyltransferase (SSAT), is responsive to antiproliferative agents. The role of SSAT in cellular responses to X-ray irradiation was examined. MATERIALS AND METHODS: Exponentially growing HeLa S3 cells were irradiated by X-rays, and mRNA levels for SSAT were measured as a function of post-irradiation time through Northern hybridization. Reverse transcription-polymerase chain reaction (RT-PCR) was used to detect alternatively spliced SSAT mRNAs. The intracellular polyamine content was measured by the o-phthalaldehyde method and the enzymatic activity of SSAT by the increased amount of acetylated spermidine after incubation. RESULTS: Not only SSAT mRNA, but also an alternatively spliced mRNA accumulated at the initial stage of growth inhibition after the first or second replication of irradiated cells. The maximum fold increase relative to the level of non-irradiated cells was 3.0-3.5 for both transcripts after 5-Gy irradiation. On the other hand, the mRNA of ornithine decarboxylase, a key enzyme of polyamine synthesis, was little influenced by X-ray treatment. Enzymatic activity of SSAT and the acetylspermidine level were elevated after X-ray irradiation. CONCLUSIONS: Activation of SSAT and the induction of alternatively spliced mRNA of the SSAT gene play an important role in regulating growth inhibition and cell death after X-ray irradiation.

Acetyltransferases↗

PubMed2004

Electromyographic and mechanomyographic estimation of motor unit activation strategy in voluntary force production.

Electromyographic and mechanomyographic estimation of motor unit activation strategy in voluntary force production. In order to determine whether electromyogram (EMG) and mechanomyogram (MMG) are suitable for the noninvasive estimation of the motor unit (MU) activation strategy, the EMG/force and MMG/force relationships were examined simultaneously during isometric ramp contractions in biceps brachii muscle. The highest mean power frequency (MPF) of the EMG, which reflects the full MU recruitment, was determined at 51% MVC. Two obvious inflection points were identified on the MMG-amplitude/force relationship that showed an initial slow increase followed by a rapid increase and a progressive decrease at higher force levels. Our results suggest that the MMG amplitude allows the estimation of the beginning of recruitment of MUs that innervate the first-twitch fibers in addition to identification of the full MU recruitment. The rate coding strategy was qualitatively reflected by the MMG-MPF/force relationship. We conclude that the MU activation strategy is estimated in more detail by the MMG than by the EMG.

Adult↗

PubMed2003

Annotation pattern of ESTs from Spodoptera frugiperda Sf9 cells and analysis of the ribosomal protein genes reveal insect-specific features and unexpectedly low codon usage bias.

MOTIVATION: A whole set of Expressed Sequence Tags (ESTs) from the Sf9 cell line of Spodoptera frugiperda is presented here for the first time. By this way we want to identify both conserved and specific genes of this pest species. We also expect from this analysis to find a class of protein sequences providing a tool to explore genomic features and phylogeny of Lepidoptera. RESULTS: The ESTs display both housekeeping as well as developmentally regulated genes, and a high percentage of sequences with unknown function. Among the identified ORFs, almost all ribosomal proteins (RPs) were found with high EST redundancy and hence sequence accuracy. The codon usage found among RP genes is in average surprisingly much less biased in Lepidoptera than in other organisms. Other Spodoptera genes also displayed a low bias, suggesting a general genome expression feature in this Lepidoptera. We also found that the L35A and L36 RP sequences, respectively, display 40 and 10 amino-acid insertions, both being present only in insects. Sequence analysis suggests that they are probably not subjected to a strong selective pressure and may be good phylogenetic markers for Lepidoptera. Most interestingly, the Lepidoptera sequences of 9 RP genes displayed a specific signature different from the canonical one. We conclude that the RP family allows valuable comparative genomics and phylogeny of Lepidoptera. AVAILABILITY: All EST sequence data are available from the private 'Spodo-Base' upon request.

Abstracting and Indexing↗

PubMed2003

Genomic sequence of a 320-kb segment of the Z chromosome of Bombyx mori containing a kettin ortholog.

The sex chromosome constitution of the silkworm, Bombyx mori, is ZW in the female and ZZ in the male. Very little molecular information is available about the Z chromosome in Lepidoptera, although the topic is interesting because of the absence of gene dosage compensation in this chromosome. We constructed a 320-kb BAC contig around the Bmkettin gene on the Z chromosome in Bombyx and determined its nucleotide sequence by the shotgun method. We found 13 novel protein-coding sequences in addition to Bmkettin. All the transposable elements detected in the region were truncated, and no LTR retrotransposons were found, in stark contrast to the situation on the W chromosome. In this 320-kb region, four genes for muscle proteins (Bmkettin, Bmtitin1, Bmtitin2, and Bmprojectin) are clustered, together with another gene (Bmmiple) on the Z chromosome in B. mori; their orthologs are also closely linked on chromosome 3 in Drosophila, suggesting a partial synteny. Real-time RT-PCR experiments demonstrated that transcripts of 13 genes of the 14 Z-linked genes found accumulated in larger amounts in males than in female moths, indicating the absence of gene dosage compensation. The implications of these findings for the evolution and function of the Z chromosome in Lepidoptera are discussed.

Amino Acid Motifs↗

PubMed2003

Reliability of the mechanomyogram detected with an accelerometer during voluntary contractions.

The accelerometer is used for mechanomyogram (MMG) recordings of muscle contractions. Although the mechanical characteristics of other MMG transducers have been determined with reference to the accelerometer, mechanical aspects of the accelerometer itself, including the weight of the transducer, have not been verified. This study was designed to reinvestigate the mechanical variable of the MMG signal detected with an accelerometer, with reference to a laser distance sensor (LDS), and then to clarify the influence of the accelerometer weight on the MMG recording during muscle contractions. The study was performed during mechanical sinusoidal vibrations and during voluntary contractions of the quadriceps muscle. Maximum differences in the amplitude spectral density functions between the LDS signal and the double integral of the accelerometer signal were approximately 4 microm. The results verified that the MMG signal from the accelerometer accurately reflected the acceleration of body surface vibration. However, the MMG signal was gradually distorted when weight was added to the accelerometer: the addition of 4.0 g (total 6.0 g, including 2.0 g of accelerometer) substantially attenuated the MMG signal. The results suggest that the appropriate weight for the accelerometer should be less than 5.0 g for measurements of the quadriceps muscle and indicate that the transducer weight must be taken into account for accurate measurement of muscles of different sizes.

Adult↗

PubMed2002

Characterization of the kynurenine 3-monooxygenase gene corresponding to the white egg 1 mutant in the silkworm Bombyx mori.

Kynurenine 3-monooxygenase (KMO, EC 1.14.13.9), which catalyzes the oxidation of kynurenine to 3-hydroxykynurenine, is involved in the synthesis of ommochrome pigments in insects. A silkworm mutant, white egg 1 ( w-1), has been shown to be deficient in this enzyme activity. The mutant is characterized morphologically by its white eyes and the fact that the females lay white eggs. To analyze the relationship between the KMO gene and the mutation, we first determined the entire sequence of a full-length 2.0-kb cDNA and examined its expression pattern in the wild type. The cDNA sequence contains one ORF encoding a polypeptide of 456 amino acids, and transcripts were detected in the larval Malpighian tubules and the pupal ovaries, but not in other tissues. Southern analysis and nucleotide sequencing showed that the KMO gene is present in a single copy and consists of ten exons distributed over a 16-kb region. Comparison of the transcripts between the wild type and mutant silkworms showed that the wild type expressed a single transcript, whereas the mutant exhibited markedly reduced amounts of two transcripts with sizes of 2.0 kb and 1.8 kb. Nucleotide sequence analysis of these mutant transcripts indicated that sequences corresponding to the ninth and tenth exons were missing. Inverse PCR and Southern analysis of the mutant gene demonstrated that the corresponding genomic region was deleted in the w-1 mutant.

Amino Acid Sequence↗