BMC Genomics 2005, 6:152 http://www.biomedcentral.com/1471-2164/6/152 175 kb of DNA. Over 5 Mb downstream of these loci are several other genes that span the human MHC region, although these are not all found adjacent to each other. In humans, MHC class I molecules are expressed on the sur- face of most cells and are recognized by CD8+ cytotoxic T cells. They are heterodimers composed of an alpha subu- nit, encoded by the MHC, and a beta subunit, or b-2 microglobulin (B2M), which maps to chromosome 15q21–q22.2. In zebrafish too, the b2m gene is found located in a different part of the genome on chromosome 13. Previously unmapped class I ZE lineage genes [20] and novel MHC class I antigen fragments have been identified on chromosome 1 and scaffold 1723. The additional functional class I lineage (mhc1ze) is also present in two other cyprinids (carp and barbus), and not linked to other class I and II genes. Analysis of the MHC class II region in zebrafish MHC class II molecules are heterodimers comprising A (alpha) and B (beta) chains that present peptides to CD4+ T cells via the endosomal pathway. One class IIA (mhc2daa) and six class IIB (mhc2dab, mhc2dbb, mhc2dcb, mhc2ddb, mhc2deb, mhc2dfb) genes have pre- viously been identified by screening a zebrafish genomic BAC library [21]. Only the mhc2dab and mhc2daa genes are known to be expressed [22]. They are closely linked and were identified on chromosome 8 (Table 2). Analysis of approximately 1 Mb of contiguous DNA surrounding the functional class II region in zebrafish demonstrates the presence of 23 flanking genes mapping to various human chromosomal locations, including two or more genes mapping to human 6q (C6orf117, LOC557721, ppil4), 12q (slc15a4, KIAA1944), 20q (rnpc2, slc12a5), 22q (slc7a4, sf3a1) and Xp (pqbp1l, t541, jarid1c). The lack of genes mapping to the human MHC, in addition to the low gene density of this region, indicates that the func- tional zebrafish class II region is the result of a transloca- tion event [12]. Mapping to chromosome 19, approximately 22 Mb telo- meric of the class I region, is the class IIB gene mhc2dcb-rs (Q95HJ7), with a similar sequence on contig NA4006. Also within this segment of DNA there are two predicted class IIA chain-encoding gene fragments, consisting of only the alpha2 domain and cytoplasmic tail-encoding parts. It is therefore unlikely that this locus is functional. Assuming that these gene fragments are not due to errors in the whole-genome assembly, it is evidence for a linkage of remnants of MHC-related class II genes with the core MHC region containing the class I peptide presenting genes. Previously identified mhc2dbb and mhc2dfb genes are pre- dicted to be located in currently unmapped contigs, NA14232 and scaffold 2399. However, mhc2deb (found in clone U08874) was not identified in the current assembly, and may be attributed to gaps in the sequence data or allelic or haplotypic differences. Several fragmented sequences resembling class IIA and/or B chain-encoding genes were also identified in contigs NA17244 and NA15003, and chromosomes 5, 15 and 20. Only contig NA6696 harbours putative complete class IIA and class IIB genes. Another gene found in this 15 kb contig resembles mhc2ddb, consisting of exons 3 and 4 only. The two TAP transporters in human, namely TAP1 and TAP2, are both located in the MHC class II region, and are closely linked to genes encoding the proteasome subunits and the class II molecules. In zebrafish, however, a TAP1- like sequence is found on chromosomes 9 and three TAP2- like genes are found in the class I region [15] on chromo- some 19. The latter, named abcb3, abcb3l1, abcb3l2, form part of the zebrafish core MHC region. The trout tap1 gene is not linked to the major class IA region either [23]. Like- wise, in Fugu, the tap1 gene is found on an isolated scaf- fold that is not linked to the main class I region [24]. Analysis of the MHC class III region in zebrafish The human class III is the most gene dense region within the MHC containing few or no pseudogenes. Unlike the class I and class II loci that are evolutionary and function- ally related [25], the class III region genes are not. The class III, however, include immune-related genes such as those encoding complement components, tumour necro- sis factors and heat shock proteins. The search for the MHC class III region in zebrafish was first initiated by the identification of the BF/C2 gene using degenerate PCR [26]. This was then followed by the identification of sev- eral, but not all, of the zebrafish homologues of the human MHC class III region genes [16]. In the zebrafish, the MHC class III genes are found distrib- uted throughout their genome (Table 3). Several zebrafish genes (zgc:63773, tnf, bat2-like, ck2b, ddah1, skiv2l, C6orf31-like, ppt2, bat3-like) map to chromosome 19, which also encompasses the largest stretch of MHC- related genes. Two other unmapped scaffolds also har- bour several class III genes: neu1, zgc:64108, bat8, zbtb12 are found in NA17767; and bf/c2, rdbp, hsp70, skiv2l are in scaffold NA17761. These are syntenic to the human MHC class III region with the conservation of both gene order and content (Figure 1). Approximately half of the human MHC class III region genes were not identified in the zebrafish assembly. Among these were the Ly6 family members, which may therefore be mammalian-specific. Alternatively, being involved in the immune response, Ly6 genes evolved more rapidly than others, and might have diverged suffi- ciently to not be recognised by sequence similarity Page 6 of 10 (page number not for citation purposes)