Сравнительный анализ генома медоносной пчелы A. M. MELLIFERA L.

There was only two sequenced genomes of the two Dipterans species Drosophila melanogaster [Adams et al., 2000] and Anopheles gambiae [Holt et al., 2002] before the recent sequencing of the honeybee Apis mellifera (Hymenoptera), Tribolium castaneum and Bombyx mori genomes (fig. 1). Hymenoptera diverged from Dipterans about 300 million years ago, and recent phylogenetic evidence implies that the Apis are the most distant group of holometabolous insects from Drosophila [Whiting, 2002; Krauss et al., 2005; Dearden et al., 2006]. Figure 1. A. A Dipteran species of the fruit fly Drosophila melanogaster. B. A Hymenopteran species of the honeybee Apis mellifera.
The honeybee genome was compared with the well-annotated, finished D. melanogaster genome. The insect D. melanogaster genome is most studied of all the genomes. Despite honeybee A. mellifera is very economically important insect a few studies of full genome its have been published [Aronstein et al., 2016; Fried, Fried, 2016; Madras–Majewska et al., 2016; Pritchard, 2016; Skonieczna, 2016; Thunman, 2016]. Therefore, comparative analysis of two genomes the A. mellifera and D. melanogaster is very interesting [Ilyasov et al., 2015; Ilyasov, 2016].
Differences between A. mellifera and D. melanogaster caused by not only the nucleotide polymorphism of the genes but also by their different epigenetical regulation. Superficially, A. mellifera development is similar to that of D. melanogaster, in that it is a holometabolous. However, A. mellifera are different in their development and biology from the D. melanogaster in a number of ways. There is a hypothesis that all the differences that are observed between A. mellifera and D. melanogaster, have occurred since their divergence. This hypothesis is confirmed by the differences observed between A. mellifera and D. melanogaster in the early stages of development [Crozier, Crozier, 1993; Whitfield et al., 2006; Weinstock et al., 2007]. Thus, A. mellifera use haplodiploidy to determine sex, a process different from that of sex determination in D. melanogaster. The adult honeybees A. mellifera has several novel evolutionary innovations not present in D. melanogaster, including poison organs and wax glands. Most important are the caste determination and labour division associated with the social nature of the honeybee [Dearden et al., 2006; Ilyasov, 2016].
The nuclear genome of the honeybees A. mellifera has 246 927 000 bp which subdivided into 16 chromosomes and containing 10 157 genes (GeneBank access AADG00000000) [Whitfield et al., 2006; Weinstock et al., 2007]. The mitochondrial genome of the honeybees has 16 343 bp which represented by a circular molecule of DNA and containing 35 genes (GeneBank access NC_001566) [Crozier, Crozier, 1993]. All chromosomes of the honeybees has different sizes: LG 1 (NC_007070) 30 000 bp contains 1669 genes (25 non coding genes); LG 2 (NC_007071) 15500 bp - 814 genes (27 non coding genes); LG 3 (NC_007072) 13200 bp - 735 genes (20 non coding genes); LG 4 (NC_007073) 12700 bp - 709 genes (46 non coding genes); LG 5 (NC_007074) 14400 bp - 874 genes (13 non coding genes); LG 6 (NC_007075) 18500 bp - 844 genes (15 non coding genes); LG 7 (NC_007076) 13200 bp - 596 genes (9 non coding genes); LG 8 (NC_007077) 13500 bp - 873 genes (33 non coding genes); LG 9 (NC_007078) 11100 bp - 584 genes (17 non coding genes); LG 10 (NC_007079) 13000 bp - 768 genes (11 non coding genes); LG 11 (NC_007080) 14700 bp - 968 genes (16 non coding genes); LG 12 (NC_007081) 11900 bp - 504 genes (14 non coding genes); LG 13 (NC_007082) 10300 bp - 418 genes (13 non coding genes); LG 14 (NC_007083) 10300 bp - 612 genes (8 non coding genes); LG 15 (NC_007084) 10200 bp - 730 genes (30 non coding genes); LG 16 (NC_007085) 7200 bp - 420 genes (26 non coding genes) (GenBank - http://www.ncbi.nlm.nih.gov, EnsemblMetazoa - http://metazoa.ensembl.org).
For comparison, the nuclear genome of the fruit flies D. melanogaster has 143 726 000 which subdivided into four chromosomes (2 large, 1 small autosomes and the X/Y sex chromosomes) and containing 17 651 genes (3 384 non coding genes) (GeneBank access GCA_000001215.4) [Adams et al., 2000]. The mitochondrial genome of the fruit flies has 19 524 bp which represented by a circular molecule of DNA and containing 37 genes (GeneBank access NC_024511.2) [Dearden et al., 2006]. All chromosomes of the fruit flies has different sizes: 2L (NT_033779.5) 23510 bp contains 3485 genes (770 non coding genes); 3L (NT_037436.4) 28110 bp contains 3453 genes (666 non coding genes); 4 (NC_004353.4) 1350 bp contains 112 genes (26 non coding genes); X (NC_004354.4) 23 540 bp contains 2661 genes (408 non coding genes); Y (NC_024512.1) 3 670 bp contains 113 genes (28 non coding genes) (GenBank - http://www.ncbi.nlm.nih.gov, EnsemblMetazoa - http://metazoa.ensembl.org).
The nuclear and mitochondrial genome of A. mellifera differ from D. melanogaster by high containing of AT-rich regions. Since the A. mellifera’s nuclear genome contains 67% and the mitochondrial genome - 85% AT whereas D. melanogaster’s nuclear genome contains 58% and the mitochondrial genome - 79% AT nucleotides [Jukes, Bhushan, 1986; Ilyasov et al., 2015].
The nuclear and mitochondrial genome of A. mellifera characterized by greater spatial heterogeneity of AT-rich areas, higher content of CpG islands and absence of the the most common families of transposones than at D. melanogaster. The genes of A. mellifera predominantly located in AT-rich areas and characterized by high content of GC nucleotides. The A and T nucleotides of AT-rich areas in protein coding genes of A. mellifera are located in second and third positions of codons predominantly [Whitfield et al., 2006; Weinstock et al., 2007].
The structure and localization of most common genes in A. mellifera differ from D. melanogaster. In the A. mellifera mitochondrial genome 11 genes of tRNA have shift position as compared with D. melanogaster. The genetic code of A. mellifera similar to D. melanogaster but two anticodons of tRNA differ (tRNALYS – TTT, tRNASER - TCT in A. mellifera and tRNALYS – CTT, tRNASER - GCT in D. melanogaster) [Crozier, Crozier, 1993].
Some nuclear genes of the A. mellifera are orthologs to the D. melanogaster genes, which has differences in sizes [Weinstock et al., 2007; Wang et al., 2014]. Thus, in A. mellifera the genes of Yellow/Major Royal Jelly Protein are larger, the genes of cuticular proteins are smaller, the genes of odorant receptors are larger, the genes of gustatory receptors are smaller, the genes of immunity are smaller, the detoxification genes are smaller than in D. melanogaster.
In the A. mellifera genes the transversions occured more frequently than transition whereas in D. melanogaster it is conversely. In the A. mellifera genes transversions occurred on third position of codons. Some genes of A. mellifera arisen as a result of evolutionary changes of the genes of common with D. melanogaster ancestors. Thus, the gene of A. mellifera encoding the major protein of royal jelly is derived from the ancient gene yellow, which presented in D. melanogaster. Many genes of A. mellifera and D. melanogaster is similar, but some genes of D. melanogaster is absent in A. mellifera. For example, in A. mellifera, the genes of WNT cell signalling pathways as HEDGEHOG (HH), TRANSFORMING GROWTH FACTOR-B (TGF-B), RECEPTOR TYROSINE KINASE (RTK), NOTCH, JANUS KINASE (JAK), SIGNAL TRANSDUCER AND ACTIVATOR OF TRANSCRIPTION (STAT) are similar with D. melanogaster. However, the genes of cell signalling systems (TERMINAL EMBRYO FATE, TRUNK, TORSO), of component of the dorso-ventral signalling system (GURKEN), of the G-protein-coupled receptor (mGluR-like) family (BOSS) are missing from the A. mellifera genome [Weinstock et al., 2007].
Some genes of the D. melanogaster has novel features in the A. mellifera. Thus, the gene of the Glucose-methanol-choline oxidoreductases family (NINAG) in A. mellifera presents as two putative NINAG-like genes, the gene of the receptor protein tyrosine kinase family (INR) in A. mellifera is duplicated; the gene of the phospholipase C family (NORPA) in A. mellifera is duplicated; the gene of the photoreceptor-cell-specific nuclear receptor family (PNR) in A. mellifera presents as three genes versus two genes in D. melanogaster; the gene of the TRPA subfamily of transient receptor potential channels family (TRPA1) are missing in D. melanogaster, but has two extra TRPA channels (GB14005 and GB16385) in A. mellifera; the gene of the ligand-gated ion channels family (NACR) in A. mellifera presents as 11 subunits instead of 10 in D. melanogaster; the gene of the ligand-gated ion channels family (NMDAR) in A. mellifera presents as 3 genes instead of 2 in D. melanogaster; the gene of the excitatory amino acid transporters family (EAAT) in A. mellifera presents as 5 genes instead of 2 in D. melanogaster [Weinstock et al., 2007].
In A. mellifera 96 homeobox domains were found in 74 genes, similar to D. melanogaster. More than 90% identity represented by homeobox genes (SEX COMBS REDUCED (SCR), ANTENNAPEDIA (ANTP), ABDOMINAL-A (ABD-A); ENGRAILED (EN), MUSCLE SEGMENT HOMEOBOX (MSH)). For the remaining A. mellifera genes, a D. melanogaster homologue is not known. This indicates that structurally homologous genes are involved in the control of A. mellifera and D. melanogaster development [Walldorf et al., 1989; Weinstock et al., 2007].
The nuclear genes of A. mellifera which responsible for circadian rhythms (CRY-M, CLK, CYC, PDP1, VRI, PER), RNA interference (RNAi) and DNA methylation (381 genes in eggs and sperm of A. mellifera with CpG methylation) have more similarity with genes of vertebrate than with genes of D. melanogaster [Toma et al., 2000; Rubin et al., 2006; Elango et al., 2009; Drewell et al., 2014]. The circadian rhythms genes TIMELESS (TIM1) and CRYPTOCHROME (DCRY) of D. melanogaster are absent in A. mellifera genome. The similarity with vertebrate may be explained by the parallel evolution of the some genes during adaptation to the environment conditions. The genome of A. mellifera contains less genes of the native immunity, of detoxification enzymes, of cuticle proteins and taste receptors compared with D. melanogaster. However, A. mellifera contains new genes associated with olfactory receptors, the processing of pollen and nectar which absent at D. melanogaster. Probably, this is due to the ecology of bees and their social organization [Dearden et al., 2006; Wallberg et al., 2014].
The rate of the evolutionary transformations of the nuclear and mitochondrial genome of A. mellifera less than in D. melanogaster. However, the genome of A. mellifera diverged more distantly from common ancestor than D. melanogaster [Crozier et al., 1989; Crozier, Crozier, 1992]. Probably, this is due to the small effective population size of A. mellifera and to low rate of the reverse mutation compared with D. melanogaster [Crozier, 1980].
Micro RNA (miRNA) of the nuclear genome of A. mellifera plays an important role in the regulation of social organization and caste differentiation via post-transcriptional regulation of gene expression. About 300 honeybee miRNAs deposited in miRBase (http://www.mirbase.org) [Ashby et al., 2016]. For example, differentially expressed miRNAs between 4-day-old queen and worker larvae of honeybees: up-regulated in queen larvae (ame-bantam, ame-let-7, ame-mir-10, ame-mir-100, ame-mir-6001-3p); equally expressed in queen larvae (ame-mir-11, ame-mir-1175, ame-mir-190, ame-mir-6065, ame-mir-989); down-regulated in queen larvae (ame-mir-13b, ame-mir-252a, ame-mir-2765-5p, ame-mir-996, ame-mir-9a) [Shi et al., 2015]. In the nuclear genome of A. mellifera found miRNA, which characterized by caste specific expression: the miRNA C5599F most expressed in the queens, C689F - in the pupaes, C5560 - in the pupaes of workers [Whitfield et al., 2006].
Thus, the genome of A. mellifera have more similarity with the vertebrate genome than D. melanogaster. The genome of A. mellifera contains less genes of the native immunity, of detoxification enzymes, of cuticle proteins and taste receptors compared with D. melanogaster. However, A. mellifera contains new genes associated with olfactory receptors, the processing of pollen and nectar, poison organs, wax glands, caste determination and labour division which absent at D. melanogaster. Probably, this is due to the ecology of bees and their social evolution. A comparative analysis of the genomes of A. mellifera and D. melanogaster using bioinformatics techniques allowed revealing the features of the structure and function of the honeybee A. mellifera genome.

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