Chromosomal location, exon/intron organization and evolution of lipocalin genes

Lipocalins exhibit low sequence similarity that contrasts with a tightly conserved folding shared by all members of this superfamily. This conserved folding can be, at least partly, accounted for by a highly conserved gene structure. The array of lipocalin genes that have so far been studied mostly in mammals indicate a large conservation of a typical seven exon/six intron arrangement. Other conserved features include a partly coding exon 1 of variable size, fixed sizes of exons 2-5 that code for an array of lipocalin-specific beta-strands and a tendency of the last exons to either fuse or expand into further exons without major changes in the length of the resulting open reading frame. The conserved exon/intron arrangement as well as a clustering of most lipocalin genes in given chromosomes of human and mouse indicate that the lipocalin genes diverged from a shared ancestor by successive rounds of duplications followed by late changes in exon arrangements.     

Human 27-kDa calbindin complementary DNA sequence. Evolutionary and functional implications

Human 27-kDa calbindin cDNA clones were selected by antibody screening from lambda gt11 brain libraries. The sequence revealed an open reading frame coding for a protein of 261 amino acids, containing four active calcium-binding domains, and two modified domains that had presumably lost their calcium-binding capability. Comparison with chick and bovine calbindins showed that the protein was highly conserved in evolution (evolutionary rate: 0.3 x 10(-9) amino acid-1 year-1) and that active and inactive domains were equally conserved. From the data we postulate that calbindin has an important physiological function involving protein--protein interactions. Comparison of calcium-binding domains from various proteins suggested that all members of the troponin C superfamily derive from a common two-domained ancestor, but that duplications leading to calbindin and to the four-domained calcium-binding proteins took place independently on different branches of the evolutionary tree. Preliminary data showed that another calcium-binding protein, homologous to calbindin, is present in the brain and encoded by a different gene.     

Maize genes encoding the small subunit of ADP-glucose pyrophosphorylase

Plant ADP-glucose pyrophosphorylase (AGP) is a heterotetrameric enzyme composed of two large and two small subunits. Here, we report the structures of the maize (Zea mays) genes encoding AGP small subunits of leaf and endosperm. Excluding exon 1, protein-encoding sequences of the two genes are nearly identical. Exon 1 coding sequences, however, possess no similarity. Introns are placed in identical positions and exhibit obvious sequence similarity. Size differences are primarily due to insertions and duplications, hallmarks of transposable element visitation. Comparison of the maize genes with other plant AGP small subunit genes leads to a number of noteworthy inferences concerning the evolution of these genes. The small subunit gene can be divided into two modules. One module, encompassing all coding information except that derived from exon 1, displays striking similarity among all genes. It is surprising that members from eudicots form one group, whereas those from cereals form a second group. This implies that the duplications giving rise to family members occurred at least twice and after the separation of eudicots and monocot cereals. One intron within this module may have had a transposon origin. A different evolutionary history is suggested for exon 1. These sequences define three distinct groups, two of which come from cereal seeds. This distinction likely has functional significance because cereal endosperm AGPs are cytosolic, whereas all other forms appear to be plastid localized. Finally, whereas barley (Hordeum vulgare) reportedly employs only one gene to encode the small subunit of the seed and leaf, maize utilizes the two genes described here.     

Evolution of the fibronectin gene. Exon structure of cell attachment domain

Genomic DNA coding for human fibronectin was identified from a human genomic library by screening with a cDNA clone that specifies the cell attachment domain in human fibronectin. Two clones which together provided more than 22 kilobase pairs of the fibronectin gene were isolated. The exons in this region correspond to approximately 40% of the coding region in the fibronectin gene. They code for the middle region of the polypeptide which consists of homologous repeating segments of about 90 amino acids called type III homologies. Nucleotide sequence of the portion of the gene corresponding to the cell attachment domain showed that the Arg-Gly-Asp-Ser cell attachment site is encoded within a 165-base pair exon. This exon, together with a 117-base pair exon codes for a homology unit. Analysis of the exon/intron organization in some of the neighboring homology units indicated a similar 2-exon structure. An exception to this pattern is that a single large exon codes for a type III homology unit that, due to alternative mRNA splicing, exists in some but not all fibronectin polypeptides. The introns separating the coding sequences for the type III homology units are located in conserved positions whereas the introns that interrupt the coding sequence within the units are in a variable position generating variations in the size of the homologous exons. This exon/intron organization suggests that the type III homology region of the fibronectin gene has evolved by a series of gene duplications of a primordial gene consisting of two exons. Specification of one of these homology units to the cell attachment domain has occurred within this exon/intron arrangement.     

Polymorphism across an exon-intron boundary in an avian Mhc class II B gene.

Twenty-three sequence haplotypes spanning the boundary of the second exon and intron of a red-winged blackbird Mhc class II B gene, Agph-DAB1, are presented. The polymorphism of the exon segment is distributed in two divergent allelic lineages which appear to be maintained by balancing selection. The silent nucleotide diversity of the exon (pi = 0.101) is more than five times that of the intron (pi = 0.018) and decays rapidly across the exon-intron boundary. Additionally, genealogical reconstruction indicates that divergence from a common ancestor in the exon sample is over four times that of the intron. The intron sequences reveal a pattern of polymorphism which is characteristic of directional selection, rather than a pattern expected from linkage to a balanced polymorphism. These results suggest that the evolutionary histories of these two adjacent regions have been disassociated by recombination or gene conversion. The estimated population recombination parameter between the exon and the intron is sufficiently high (4NeC = 8.545) to explain the homogenization of intron sequences. Compatibility analyses estimate that these events primarily occur from the exon-intron boundary to about 20-30 bases into the intron. Additionally, the observation that divergent exon alleles share identical intron sequence supports the conclusion of disassociation of exon and intron evolutionary histories by recombination.     

动物肌动蛋白基因中内含子的来源及存在意义的探讨

对动物界演化过程中肌动蛋白家族内含子插入位置分布的演化规律作了分析,并对相同插入位置的内含子序列按同亚型和不同亚型作了比较。结果得出:从整个肌动蛋白家族的外显子序列高度保守性推断整个肌动蛋白家族可能是从共同的祖先蛋白进化而来的;从同亚型肌动蛋白内含子序列的类似性随进化距离而变化,但在短进化距离的物种间,类似性都较高,不同亚型肌动蛋白内含子序列的类似性都较低,即使是同一物种(如人),类似性也远低于同亚型但进化距离较近的物种,由此可推断,同亚型肌动蛋白的内含子序列可能从共同祖先进化,不同亚型肌动蛋白的内含子序列从不同祖先进化,综上推断可导出内含子可能是在蛋白异化过程中获得的:还发现内含子在肌动蛋白家族编码基因中位置的分布随进化方向不同而逐步形成两种截然不同的模式,由此提出了内含子的位置分布与动物演化方向之间可能具有某种必然联系,为内含子的存在提出了某种依据。     

The human genes for calbindin 27 and 29 kDa proteins are located on chromosomes 8 and 16, respectively

Genomic clones coding for the brain calcium-binding protein, calbindin 29 kDa, were isolated from a human library. A fragment containing exon 2 was used as a probe to investigate the presence of the gene in human x rodent somatic cell hybrids. The gene was unambiguously assigned to chromosome 16. The closely-related calbindin 27 kDa gene was previously assigned to chromosome 8. These two genes, deriving from a common ancestor, thus appear to have been separated during vertebrate evolution.     

Evolutionary lineages of RT1.Ba in the Australian Rattus

In this study, the evolutionary history of the variable second exon of RT1.Ba and its adjoining intron b are compared across a number of species and subspecies of the Australian RATTUS: Three lineages are identified in the second intron across a range of Rattus species. Two of these lineages, separated by the insertion of a probable rodent short interspersed nucleotide element and by point mutations outside the indel region, are both found in each of the major clades of the endemic Australian RATTUS: This pattern of ancestral polymorphism is reflected in the adjoining exon 2 sequences, although phylogenetic constraints confirm that the clustering is not identical to that of the associated intron sequences. In addition, the coding sequences show evidence of the retention of ancestral polymorphism, with identical exon sequences found in two divergent species, and some indication of gene conversion detected for the exon sequences.     

Prevalence of intron gain over intron loss in the evolution of paralogous gene families

The mechanisms and evolutionary dynamics of intron insertion and loss in eukaryotic genes remain poorly understood. Reconstruction of parsimonious scenarios of gene structure evolution in paralogous gene families in animals and plants revealed numerous gains and losses of introns. In all analyzed lineages, the number of acquired new introns was substantially greater than the number of lost ancestral introns. This trend held even for lineages in which vertical evolution of genes involved more intron losses than gains, suggesting that gene duplication boosts intron insertion. However, dating gene duplications and the associated intron gains and losses based on the molecular clock assumption showed that very few, if any, introns were gained during the last ~100 million years of animal and plant evolution, in agreement with previous conclusions reached through analysis of orthologous gene sets. These results are generally compatible with the emerging notion of intensive insertion and loss of introns during transitional epochs in contrast to the relative quiet of the intervening evolutionary spans.     

Dynamic Insertion–Deletion of Introns in Deuterostome EF-1α Genes

To test the validity of intron–exon structure as a phylogenetic marker, the intron–exon structure of EF-1α genes was investigated for starfish, acornworms, ascidians, larvaceans, and amphioxus and compared with that of vertebrates. Of the 11 distinct intron insertion sites found within the coding regions of the deuterostome EF-1α genes, 7 are shared by several taxa, while the remainder are unique to certain taxa. Examination of the shared introns of the deuterostome EF-1α gene revealed that independent intron loss or intron insertion must have occurred in separate lineages of the deuterostome taxa. Maximum parsimony analysis of the intron–exon data matrix recovered five parsimonious trees (consistency index = 0.867). From this result, we concluded that the intron–exon structure of deuterostome EF-1α has evolved more dynamically than previously thought, rendering it unsuitable as a phylogenetic marker. We also reconstructed an evolutionary history of intron insertion–deletion events on the deuterostome phylogeny, based on several molecular phylogenetic studies. These analyses revealed that the deuterostome EF-1α gene has lost individual introns more frequently than all introns simultaneously.     

Conserved exon-intron organization in two different caerulein precursor genes of Xenopus laevis. Additional detection of an exon potentially coding for a new peptide

From a genomic library of Xenopus laevis, two genes coding for different preprocaeruleins have been isolated and sequenced. These correspond to the type I and type III precursors analyzed previously at the cDNA level [Richter, K., Egger, R. and Kreil, G. (1986) J. Biol. Chem. 261, 3676-3680]. The type III gene comprises eight exons; the type I apparently contains eight exons as well, of which six have been sequenced. The genetic information for the dekapeptide caerulein is present on small exons of 45 base pairs. The two genes are highly homologous in their 5'-flanking region, the exon/intron boundaries, and long stretches of intron sequences. A possible scheme for the evolution of this small family of genes through exon and gene duplications is presented. In the type I gene, in place of one of the caerulein exons, a potential exon with conserved splice sites was discovered. If expressed in some frog cells, this exon would code for a new peptide 60% homologous to caerulein.     

Transfer of rpl22 to the nucleus greatly preceded its loss from the chloroplast and involved the gain of an intron.

Most chloroplast and mitochondrial proteins are encoded by nuclear genes that once resided in the organellar genomes. Transfer of most of these genes appears to have occurred soon after the endosymbiotic origin of organelles, and so little is known about the process. Our efforts to understand how chloroplast genes are functionally transferred to the nuclear genome have led us to discover the most recent evolutionary gene transfer yet described. The gene rpl22, encoding chloroplast ribosomal protein CL22, is present in the chloroplast genome of all plants examined except legumes, while a functional copy of rpl22 is located in the nucleus of the legume pea. The nuclear rpl22 gene has acquired two additional domains relative to its chloroplast ancestor: an exon encoding a putative N-terminal transit peptide, followed by an intron which separates this first exon from the evolutionarily conserved, chloroplast-derived portion of the gene. This gene structure suggests that the transferred region may have acquired its transit peptide by a form of exon shuffling. Surprisingly, phylogenetic analysis shows that rpl22 was transferred to the nucleus in a common ancestor of all flowering plants, at least 100 million years preceding its loss from the legume chloroplast lineage.     

Rapid changes in the exon/intron structure of a mammalian thrombin inhibitor gene

The genomic organization of the heparin cofactor II (HCII) gene from rat and mouse was investigated and compared with their human counterpart. The genes share a common core structure consisting of five exons interrupted by four introns, but the mouse and rat gene reveal individual additional features. A unique differentially spliced exon is present in the 5'-untranslated region of the rat gene, which most probably has arisen de novo by point mutations in intronic sequences of the ancestor gene. In the mouse HCII gene, a novel intron/exon boundary has been created due to the presence of an additional DNA segment, which simultaneously provides a 3'-splice site and a polypyrimidine stretch leading to an alternatively used exon of increased size. Our data suggest that, in contrast to most other mammalian genes, the exon/intron pattern of the gene coding for HCII is in dynamic evolution.     

Gene relocations within chloroplast genomes of Jasminum and Menodora (Oleaceae) are due to multiple, overlapping inversions

The chloroplast (cp) DNA sequence of Jasminum nudiflorum (Oleaceae-Jasmineae) is completed and compared with the large single-copy region sequences from 6 related species. The cp genomes of the tribe Jasmineae (Jasminum and Menodora) show several distinctive rearrangements, including inversions, gene duplications, insertions, inverted repeat expansions, and gene and intron losses. The ycf4-psaI region in Jasminum section Primulina was relocated as a result of 2 overlapping inversions of 21,169 and 18,414 bp. The 1st, larger inversion is shared by all members of the Jasmineae indicating that it occurred in the common ancestor of the tribe. Similar rearrangements were also identified in the cp genome of Menodora. In this case, 2 fragments including ycf4 and rps4-trnS-ycf3 genes were moved by 2 additional inversions of 14 and 59 kb that are unique to Menodora. Other rearrangements in the Oleaceae are confined to certain regions of the Jasminum and Menodora cp genomes, including the presence of highly repeated sequences and duplications of coding and noncoding sequences that are inserted into clpP and between rbcL and psaI. These insertions are correlated with the loss of 2 introns in clpP and a serial loss of segments of accD. The loss of the accD gene and clpP introns in both the monocot family Poaceae and the eudicot family Oleaceae are clearly independent evolutionary events. However, their genome organization is surprisingly similar despite the distant relationship of these 2 angiosperm families.     

Characterization and evolution of napin-encoding genes in radish and related crucifers

Three cDNA clones, encoding napin storage proteins from radish, were isolated and sequenced. They fall into two classes differing in the size of the primary translation product. Sequences of the two classes are very well conserved and they display an organization very similar to that of the homologous genes from rapeseed and Arabidopsis which have previously been described. On the basis of hybridization intensity and the number of restriction fragments, we estimate that the radish napin multigene family is represented by eight to twelve members. The use of probes specific to each subfamily demonstrates that they contribute to a similar extent to the production of napin mRNA. Analysis of the sequence data suggests that the napin ancestral genes are probably derived from successive duplication and divergence of a protogene. Comparing other available napin sequences with those of radish reveals intriguing features. Comparison of the coding sequences shows that the homology between the radish and rapeseed sequences is much higher than that between each of the four members of the Arabidopsis gene family. This would suggest that the duplications which gave rise to the different members occurred independently in the two groups of species after separation of Arabidopsis from the Brassica lineage. However, similar comparison carried out on the 3' -noncoding sequences does not support this hypothesis, but shows that slightly different duplicated genes probably already existed in the common ancestor to the three genera. This paradox can be resolved by assuming that, within each genus, coding sequences for napin-encoding genes have been considerably homogenized as a result of concerted evolution.     

The complete intron/exon structure of Ephydatia mülleri fibrillar collagen gene suggests a mechanism for the evolution of an ancestral gene module

We have completed the analysis of a genomic clone, G238, that contains most of the coding region of the sponge COLF1 fibrillar collagen gene. The main triple helical domain is encoded by 31 exons. Except for the 5 junction exon and the two last 3 exons (126 and 18 base pairs), all these exons are related to a 54-bp unit and begin with an intact glycine codon. A good correlation can be made between this sponge gene and a vertebrate fibrillar collagen gene, revealing the high conservation of the members of this family during evolution. The reconstitution of an ancestral collagen gene can be made by considering all the exon/intron junctions of these genes. We suggest that such an ancestral gene arose from multiple duplications of a 54-bp exon and a (54 + 45)-bp module.Abbreviations used bp base pair(s) - kb kilobase(s) - C-protease the enzyme that cleaves the carboxyl-terminal propeptide