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Genome-wide survey and characterization of transcription factors in the silk gland of the silkworm, Bombyx mori

ABSTRACT: Transcription factors (TFs) are key proteins that modulate gene transcription and thereby lead to changes in the gene expression profile and the subsequent alteration of cellular functions. In the silk gland (SG) of silkworm Bombyx mori, an important silk-producing insect, TFs are of vital importance in the regulation of silk protein synthesis in this organ. However, which TFs exist and express in the SG remains largely unknown. Here, we report the large-scale identification of TFs in the SG based on available full-length transcript sequences and the most recent version of silkworm genome data. In total, 348 candidate TFs were identified by strict filtration and were classified into 56 TF families. Chromosomal distribution, motif composition, and phylogenetic relationship analyses revealed the typical characteristics of these TFs. In addition, the expression patterns of 348 TFs in various tissues of B. mori, especially the SG of fourth-molt (4LM) and day-3 and day-4 fifth-instar (5L3D and 5L4D) larvae, were investigated based on public RNA-seq data and gene microarray data, followed by spatiotemporal verification of TF expression levels by quantitative real-time PCR (qRT-PCR). This report describes the first comprehensive analysis of TFs in the B. mori SG. The results can serve as a baseline for further studies of the roles of TFs in the B. mori SG.

SUBMITTER: Ma Y

PROVIDER: S-EPMC8584736 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:To identify functions that distinguish the posterior and median cells producing fibroin and sericin in the silk gland of Bombyx mori, serial analysis of gene expression (SAGE) profiles from both silk gland regions were analyzed and compared. The construction of a B. mori reference tag collection extracted from a set of 38000 Bombyx EST sequenced from the 3’ side, helped annotating the SAGE libraries. Most of the tags appeared at similar relative concentration in the two libraries except for those corresponding to silk proteins that were found region-specific and highly abundant. Strikingly, besides tags from silk protein mRNAs, 19 tags were found in the class of high abundance in the median cell library, which were absent in the posterior cell tag collection. Except tags from SP1 mRNA, no PSG specific tags were found in the same class of abundance. The analysis of MSG-specific different transcripts led to suggest that middle silk gland cell realizes more diversified functions as those already known, of synthesis and secretion of the silk sericins. EST libraries from 11 silkworm tissues were 3’-sequenced to ensure the identification of the most terminal tag. 37,920 sequences were analyzed on ABI 3700 or 3730XL sequencers. Electrophoregrams were processed with KB Basecaller (3730XL traces) or with PHRED (3700 traces) to obtain the .phd files from which were extracted text sequences and their corresponding quality files in Fasta format (Phd2Fasta, Green and Ewing, 1995; 2002). Vector sequences and bad quality regions were removed with an home made software after identification by Lucy (Chou and Holmes, 2001). Chimera were removed by an home made software and retrotransposon sequences were masked by RepeatMasker (http://repeatmasker.org). The cleaned sequences were clusterized with TGICL package (TIGR) and assembled into contigs with CAP3 (Huang and Madan, 1999). Contigs were identified with Blastn and Blastx on GenBank and SwissProt/TREMBL, respectively. Some clusters, splitted by CAP3 procedure, have been regrouped on the basis of Blastx identity. Identitag (Keime et al., 2004) was used to extract all possible tags (forward and reverse) to create a reference database. Moreover, a quality index is attached to each tag depending to the presense of poly-A, polyadenylation signal and its proximity to 3-prime extremity in the original mRNA sequence. This database was supplemented with the tags extracted from public B. mori sequences (GenBank, Silkbase) with the same software. Tags were extracted from MSG and PSG libraries (from 2304 and 3072 sequenced inserts respectively) with the Sagenhaft software (Beissbarth et al., 2004) then identified and compared with Identitag. The assessment of significant differences among the two libraries was performed by using the Z-test used for comparison of SAGE libraries of different size (Kal et al., 1999). For graphic purpose and to avoid division by zero we used a tag value of 1 for tags that were not detected in MSG or PSG libraries. Since the two SAGE libraries showed up different TAG number, we used the Z-test for comparing the two mRNA populations.

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Genome-wide survey and characterization of transcription factors in the silk gland of the silkworm, Bombyx mori

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