## 1 Introduction  
Alternative splicing of pre-mRNA is an essential mechanism in the regulation of differential gene expression that can produce functionally distinct proteins from a single gene based on the developmental or physiological state of the cells in a multicellular organism [1,2]. Recent studies estimate that 90% of human genes are alternatively spliced [3], and several thousand different mRNA isoforms can be produced from a single gene. Although many examples describe how alternative splicing regulates gene expression, the mechanisms involved are less well understood [4, 5, 6, 7, 8]. Alternative splicing is thought to be regulated by the interaction of splicing factors and splicing enhancers (or silencers). Alternative splicing regulatory mechanisms have been investigated, structural models of spliceosomes have been proposed, and many RNA regulatory elements have been characterized; however, the emerging complexity of alternative splicing regulation suggests that these approaches have not sufficiently described how alternative splicing is regulated. Recent provocative studies point to a key function of chromatin structure and histone modification in alternative splicing regulation [9, 10, 11]. For example, the nucleosome occupancy level was lower in cassette exons than in constitutively spliced exons [12, 13, 14]. H3K36 me3 and H3K9 ac were related to the exon-skipping event of NCAM [15] and the levels of H3K36 me3 differed in mutually exclusive exons of several genes among different cell types. Moreover, a genome-wide study across different species revealed that H3K36 me3 was depleted in skipped exons [13, 16]. Other histone modifications such as H3K4 me1, H3K4 me3, H3K27 me3, and H3K9 me1 have been associated with the alternative splicing events of FGFR2 [17]. H3K4 me3 was suggested to affect the alternative splicing events of CHD1 [18], while H3K9 me3 was associated with multiple exon skipping of CD44 [19]. A recent genome-wide chromatin immunoprecipitation (ChIP)-seq analysis of histone H3 methylation in mammals revealed that alternative exons are preferentially marked with H3K4 me1, H3K27 me3, and H3K79 me2, while being marked with H3K4 me2, H3K4 me3, and H3K36 me3 at significantly lower levels [20]. In the silkworm _Bombyx mori_, the chromosomal sex determination mechanism is distinct from that of _Drosophila melanogaster_, with female (ZW) being the heterogametic sex and male (ZZ) the homogametic sex. The female sex in _B. mori_ is determined by the presence of a dominant feminizing factor, _Feminizer_ (_Fem_), on the W chromosome [21]. Note that no sex-specific regulatory _Sxl_ homolog has been isolated from _B. mori_[22], and no _tra_ homolog has been found in the _Bombyx_ genome [23]. Despite these differences, a _B. mori dsx_ homolog (_Bmdsk_) has been implicated in sex determination [24]. The primary transcript of the _Bmdsk_ gene is alternatively spliced in males and females to yield sex-specific mRNAs that encode male-specific (BmDSXM) and female-specific (BmDSXF) polypeptides [25]. We found that unlike _Drosophila dsx_, the _Bmdsk_ female exon is devoid of putative TRA/TRA-2 binding sites [25]. Instead, the splicing inhibitor BmPSI and a _B. mori_ insulin-like growth factor II mRNA-binding protein (Imp) regulate male-specific splicing of _Bmdsk_[26, 27]. _Imp_ is localized on the Z chromosome and is expressed in a male-specific manner in various tissues. In male cells, the male-specific _Imp_ mRNA is formed as a result of the inclusion of exon 8 and the promoter-distpoly(A) site choice, whereas non-sex-specific polyadenylation occurs at the promoter-proximal poly(A) site downstream of exon 7 [28]. The molecular mechanisms underlying the sex-specific splicing regulation of this gene remain unclear. To verify the link between histone methylation and alternative RNA processing in _Imp_ mRNA production, we investigated the effects of RNAi-mediated knockdown of several histone methyltransferases (HMTases) on sex-specific mRNA expression of _Imp_.