BMC Genomics 2004, 5:69 http://www.biomedcentral.com/1471-2164/5/69 The CDK10/11 family In this group, orthologs of CDK10 are found only in human and Drosophila, while CDK11 occurs in human, Drosophila and Caenorhabditis. Three putative CDK11 orthologs were found in plants (two from Arabidopsis and one from Oryza). CDK10 has been implicated in the regu- lation of the G2/M phase of the cell cycle [52], but a cyclin partner has yet to be defined. Only one protein associated with CDK10, ETS2 transcription factor, has been identi- fied so far, suggesting a link to transcription [9]. CDK11 associates with cyclin L as a partner, and is a proposed component of a signaling pathway that helps to coordi- nate transcription and RNA-processing events [10-13]. The close relationship between the CDK10 and CDK11 may reflect evolutionary and/or mechanistic similarities, but neither kinase family has been well characterized functionally. In addition, BC18H10 from S. pombe and PfCRK1 from Plasmodium show close relationships to the CDK10/11 family, but no function has yet been deter- mined for these kinases either. Conclusions The apparent co-evolution between the CTD and certain CTD-specific kinases suggests an explanation for strong stabilizing selection on CTD structure in some eukaryotes, and its complete degeneration in others. Based on the genomes examined in this study, either the origins of CDK7 and CDK8 in an unknown ancestor of the "CTD- clade," or the canalization of reversible phosphorylation of the CTD in some eukaryotic groups but not others, could account for the variation seen in RPB1 C-terminal structure. In either case, once thoroughly "locked" into RNAP II function, the CTD must have recruited other tran- scription and processing related proteins into a growing machinery of the "transcriptosome" [53]. Our results sug- gest that was the case for several CDKs that clearly predate the canalization of CTD-based RNAP II transcription; fur- ther genomic analyses are underway to look for other pro- tein-protein interactions that could be responsible for strong evolutionary conservation of the CTD in members of the "CTD-clade." This work also provides a new perspective on the overall evolution CDKs and evolutionary relationships among kinase families. Our combined genomic and phylogenetic analyses suggest that transcription-related kinases origi- nated later than cell cycle-related CDKs. Finally, our results point to potential functions for a variety of previ- ously uncharacterized kinases, based on their apparent orthologous relationships to defined CDKs. Additional completed genomes, particularly those from broadly diverse protists (especially non-parasitic forms), will be critical to address these questions further. Such compara- tive analyses will be invaluable in helping to guide exper- imental studies, which ultimately are required to verify the functional properties of each putative CDK. Methods Identification and alignment of protein sequences Representatives of all previously identified CDKs from budding yeast and human were obtained from Genbank, and used as probes in TBlastN and PSI-Blast [54] against the National Center for Biotechnology Information (NCBI), and additional specific complete genome data- bases, with an absolute cut-off of E<0.001. To confirm the identities of putative CDKs detected by the TBlastN, each identified sequence was used as a query in reciprocal Blast searches, to verify that it retrieved the original query sequences, and global sequence alignments were per- formed to confirm putative homologies to CDKs, accord- ing to the CDC-related kinase characterized motifs that use CDK2 as the model [55]. Initially, a number of inferred protein sequences were grouped into six subsets according to clear similarities to specific CDK family orthologs. These subgroups first were aligned in CLUSTAL X [56], and the resulting sub-align- ments then were aligned with each other and adjusted through visual inspection and comparison to the kinase alignment of Liu and Kipreos (2000) [32]. Regions that could not be aligned reliably were excluded from subse- quent phylogenetic analysis. The resulting alignment included 233 positions including gaps (See additional data file 2 and 3 for the original and final aligned matrices used in this study). Phylogenetic analysis Maximum-likelihood (ML) estimates of substitution parameters were made with the program TREEPUZZLE-50 [57] assuming a mixed model for variation among sites, with one category for invariable sites and a four-category discrete approximation to Γ-distribution, and the JTT weighting matrix for probability of change among amino acids. Further analyses were performed in MrBayes 3.0 b4 [58] using metropolis-coupled Markov chain Monte Carlo analysis. Four simultaneous Markov chains were run, also under an invariant + Γ rate model and a JTT substitution matrix. Four chains, one heated, were run for 500,000 generations, beginning with random a priori trees. Trees were sampled from the posterior probability distribution every 100 generations. The empirical burn-in required for likelihoods to converge was less than 100,000 genera- tions; an additional 400,000 generations were run and the first 100,000 were excluded from analysis of Bayesian pos- terior probabilities. Thus, a total of 4,000 trees were exam- ined to determine the 50% majority-rule consensus tree and Bayesian support values. In addition, 1000 distance (PROTDIST + NEIGHBOR) bootstrap replicates were Page 11 of 13 (page number not for citation purposes)