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Molecular characterization and genetic origin of the Brassica napus acetohydroxyacid synthase multigene family.

The Brassica napus rapeseed cultivar Topas contains an acetohydroxyacid synthase (AHAS) multigene family consisting of five members (AHAS 1-5). DNA sequence analysis indicate that AHAS1 and AHAS3 share extensive homology. They probably encode the AHAS enzymes essential for plant growth and development. AHAS2 has diverged significantly from AHAS1 and AHAS3 and has unique features in the coding region of the mature polypeptide, transit peptide and upstream non-coding DNA, which raises the possibility that it has a distinct function. AHAS4 and AHAS5 have interrupted coding regions and may be defective. The complexity of the AHAS multigene family in the allotetraploid species B. napus is much greater than reported for Arabidopsis thaliana and Nicotiana tabacum. Analysis of the presumptive progenitor diploid species B. campestris and B. oleracea indicated that AHAS2, AHAS3 and AHAS4 originate from the A genome, whereas AHAS1 and AHAS5 originate from the C genome. Further variation within each of the AHAS genes in these species was found.

Acetolactate Synthase↗

Organization, evolution and expression of a multigene family encoding putative members of the odourant binding protein family in the medfly Ceratitis capitata.

A multigene family encoding male specific serum polypeptides (MSSPs) that show significant structural similarity to the family of insect odourant binding proteins, has been characterized in the medfly Ceratitis capitata. This family comprises seven members classified in three subgroups, MSSP-alpha, MSSP-beta and MSSP-gamma. The genes of subgroups alpha and beta are clustered in tandem in a 35-kb genomic region, and present an exceptionally high degree of similarity not only in their coding but also in the surrounding regions, while the genes of the gamma subgroup are drastically divergent. Although MSSPs are predominantly expressed in the male fat body, detailed expression studies suggest that individual members of this family are expressed in a distinct sex- and tissue-specific manner.

Amino Acid Sequence↗

Anaplasma marginale major surface protein 3 is encoded by a polymorphic, multigene family.

The immunodominant surface protein, MSP3, is structurally and antigenically polymorphic among strains of Anaplasma marginale. In this study we show that a polymorphic multigene family is at least partially responsible for the variation seen in MSP3. The A. marginale msp3 gene msp3-12 was cloned and expressed in Escherichia coli. With msp3-12 as a probe, multiple, partially homologous gene copies were identified in the genomes of three A. marginale strains. These copies were widely distributed throughout the chromosome. Sequence analysis of three unique msp3 genes, msp3-12, msp3-11, and msp3-19, revealed both conserved and variant regions within the open reading frames. Importantly, msp3 contains amino acid blocks related to another polymorphic multigene family product, MSP2. These data, in conjunction with data presented in previous studies, suggest that multigene families are used to vary important antigenic surface proteins of A. marginale. These findings may provide a basis for studying antigenic variation of the organism in persistently infected carrier cattle.

Amino Acid Sequence↗

An integrated approach for the comparative analysis of a multigene family: the nicotianamine synthase genes of barley.

Recent genomic projects reveal that about half of the gene repertoire in plant genomes is made up by multigene families. In this paper, a set of structural and phylogenetic analyses have been applied to compare the differently sized nicotianamine synthase (NAS) gene families in barley and rice. Nicotianamine acts as a chelator of iron and other heavy metals and plays a key role in uptake, phloem transport and cytoplasmic distribution of iron, challenging efforts for the breeding of iron-efficient crop plants. Nine barley NAS genes have been mapped, and co-linearity of flanking genes in barley and rice was determined. The combined analyses reveal that the NAS multigene family members in barley originated through at least one duplication event that occurred before the divergence of rice and barley. Additional duplications appear to have occurred within each of the species. Although we detected no evidence for positive selection of recently duplicated genes within species, codon-based tests revealed evidence for positive selection having contributed to the divergence of some amino acids. The integrated comparative and phylogenetic analysis improved our current view of NAS gene family evolution, might facilitate the functional characterization of individual members and is applicable to other multigene families.

Alkyl and Aryl Transferases↗

The Py235 proteins: glimpses into the versatility of a malaria multigene family.

Py235 rhoptry proteins, encoded by a multigene family of the rodent malaria parasite Plasmodium yoelii, combine a functional role in invasion with one of immune evasion, and have homologues in malaria parasites of humans. Investigations of Py235 are summarised and the perspectives for dissecting the molecular and biological mechanisms underlying these crucial phenomena are discussed.

Animals↗

Identification and genetic mapping of 151 dispersed members of 16 ribosomal protein multigene families in the mouse.

More than 150 individual members of 16 ribosomal protein multigene families were identified as DNA restriction fragments and genetically mapped. The ribosomal protein gene-related sequences are widely dispersed throughout the mouse genome. Map positions were determined by analysis of 144 progeny mice from both an interspecific (C57BL/6J x SPRET/Ei)F1 x SPRET/Ei and an intersubspecific (C57BL/6J x CAST/Ei)F1 x C57BL/6J backcross. In addition, 30 members of the multigene families encoding PGK1 ODC, and TPI, including five new loci for ODC and one new locus for TPI, were characterized and mapped. Interspecific backcross linkage data for 29 nonecotropic murine leukemia retroviruses endogenous to C57BL/6J mice are also reported. Transmission ratio distortions and recombination frequencies are compared between the two backcrosses.

Animals↗

Interchromosomal biased gene conversion, mutation and selection in a multigene family.

A mathematical model of the effects of interchromosomal biased gene conversion, mutation and natural selection on a multigene family is developed and analyzed. The model assumes two allelic states at each of n loci. The effects of genetic drift are ignored. The model is developed under the assumption of no recombination, but the analysis shows that, at equilibrium, there is no linkage disequilibrium, which implies that the conclusions are valid for arbitrary recombination among loci. At equilibrium, the balance between mutation, gene conversion and selection depends on the ratio of the mutation rates to the quantity [s + g(2 alpha - 1)/n], where s is the increment or decrement in relative fitness with each additional copy of one of the alleles, g is the conversion rate, and alpha is a measure of the bias in favor of one of the alleles. When this quantity is large relative to the mutation rates, the allele that has the net advantage, combining the effects of selection and conversion, will be nearly fixed in the multigene family. A comparison of these results with those from a comparable model of intrachromosomal biased conversion shows that biased interchromosomal conversion leads to approximately the same equilibrium copy number as does intrachromosomal conversion of the same strength. Interchromosomal conversion is much more effective in causing the substitution of one allele by another. The relative frequencies of interchromosomal and intrachromosomal conversion is indicated by the extent of the linkage disequilibrium among the loci in a multigene family.

Alleles↗

Characterization of the complete transcriptionally active ehrlichia chaffeensis 28 kDa outer membrane protein multigene family.

The 28kDa outer membrane proteins (P28) of Ehrlichia chaffeensis are encoded by a multigene family. The purpose of this study was to determine all the p28 gene sequences and their transcriptional activities. There were 21 members of the p28 multigene family located in a 23kb DNA fragment in the genome of E. chaffeensis. The p28 genes each contained 816-903 nucleotides with intergenic spaces of 10-605 nucleotides. All the genes were complete and were predicted to have a signal sequence. The molecular masses of the mature proteins were predicted to be 28-32kDa. The amino acid sequence identity of the P28 proteins was 20-83%. Ten p28 genes were investigated for transcriptional activity by using RT-PCR amplification of mRNA. Six of 10 tested p28 genes were actively transcribed in cell-culture grown E. chaffeensis. RT-PCR also indicated that each of the p28 genes was monocistronic. These results suggest that the p28 genes are active genes and encode polymorphic forms of the P28 proteins. The P28s were divergent among isolates of E. chaffeensis also. The large repertoire of the p28 genes in a single ehrlichial organism and antigenic diversity of the P28 among the isolates of E. chaffeensis suggest that P28s may be involved in immune avoidance.

Amino Acid Sequence↗

Classification and characterization of the rice alpha-amylase multigene family.

To establish the size and organization of the rice alpha-amylase multigene family, we have isolated 30 alpha-amylase clones from three independent genomic libraries. Partial characterization of these clones indicates that they fall into 5 hybridization groups containing a total of 10 genes. Two clones belonging to the Group 3 hybridization class have more than one gene per cloned fragment. The nucleotide sequence of one clone from Group 1, lambda OSg2, was determined and compared to other known cereal alpha-amylase sequences revealing that lambda OSg2 is the genomic analog of the rice cDNA clone, pOS103. The rice alpha-amylase genes in Group 1 are analogous to the alpha-Amy1 genes in barley and wheat. lambda OSg2 contains sequence motifs common to most actively transcribed genes in plants. Two consensus sequences, TAACAAGA and TATCCAT, were found in the 5' flanking regions of alpha-amylase genes of rice, barley and wheat. The former sequence may be specific to alpha-amylase gene while the latter sequence may be related to a 'CATC' box found in many plant genes. Another sequence called the pyrimidine box (TCCTTTTTC) was found in the alpha-amylase genes as well as other genes regulated by gibberellic acid (GA). Comparisons based on amino acid sequence alignment revealed that the multigene families in rice, barley and wheat shared a common ancestor which contained three introns. Some of the descendants of the progenitor alpha-amylase gene appear to have lost the middle intron while others maintain all three introns.

Amino Acid Sequence↗

The HSP70 multigene family of Caenorhabditis elegans.

1. The heat shock response of the nematode Caenorhabditis elegans has been characterized. 2. There are at least nine genes in the hsp70 multigene family of C. elegans. 3. Five of the hsp70 genes have been characterized and assigned to one of at least three hsp70 gene subfamilies. One of the subfamilies consists of an hsp70 protein that has the potential to be translocated into the endoplasmic reticulum and another subfamily consists of a protein that has the potential to be translocated into the mitochondria. 4. The C. elegans hsp70 multigene family has several unique characteristics including introns in the heat inducible hsp70 genes, at least one trans-spliced hsp70 mRNA and two grp78 related genes, one of which is highly heat inducible. 5. The identification and characterization of C. elegans hsp70 multigene family is the basis for a genetic characterization of the regulation and function of a gene family during the development of a multicellular eukaryote.

Animals↗

A multiplex RT-PCR assay for analysis of relative transcript levels of different members of multigene families: application to Arabidopsis calmodulin gene family.

The high degree of conservation of nucleotide sequences among different members of a multigene family poses problems in analysis of expression patterns governed by each member of the gene family. In this report we describe a simple, semi-quantitative and single tube multiplex RT-PCR assay for simultaneous and relative expression analysis with an application to all the six members of Arabidopsis calmodulin multigene family. In the multiplex primer set, individual gene specific primers were derived from 3'-untranslated region of the genes and a single common primer from the conserved exonic region. Transcriptional activation of all the members of the calmodulin gene family in response to touch was monitored. The results demonstrate that two of the genes are not regulated by touch; however, the other four that are induced by touch show a differential response including their kinetics of induction.

Arabidopsis↗

Identification and genetic regulation of the chalcone synthase multigene family in pea.

Chalcone synthase (CHS) is a key enzyme in the biosynthesis of diverse flavonoids involved in disease resistance, nodulation, and pigmentation in pea. We describe a multigene family encoding CHS and the effects of two regulatory loci, a and a2, on the pattern of expression of three of its member genes. Two of the genes, CHS1 and CHS3, are expressed in both petal and root tissue, whereas expression of a third gene, CHS2, is detected only in roots. The products encoded by the a and a2 loci are required for the expression of the CHS1 gene and for wild-type levels of expression of the CHS3 gene in petal tissue. In root tissue, all three CHS genes are expressed and induced by CuCl2 regardless of the genotype at the a and a2 loci. These results show that the various members of the CHS multigene family interact in diverse ways with multiple genetic signals in the plant, providing a basis for the differential expression of these genes. Spatially specific genetic regulation of distinct members of a multigene family has been clearly demonstrated.

Acyltransferases↗

Characterization of the rbcS multigene family in wheat: subfamily classification, determination of chromosomal location and evolutionary analysis.

To elucidate the evolution of a multigene family in plants, nucleotide sequences of members of the rbcS multigene family encoding the small subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase in wheat were determined. Five genomic clones containing rbcS sequences were isolated. The known members of the rbcS family of common wheat, including the clones reported here, were categorized into two Classes (I and II), which, in turn, were classified into three (A, B and E) and two (C and D) subtypes, respectively. The sequences of the intron and the 3' flanking region were divergent between the classes but not among the types within a class. The differences among the types within a class were caused by large indels (about 100-350 bp) in the intron. The members of Classes I and II were located on homeologous group 2 and 5 chromosomes, respectively. The Southern hybridization data indicated that all types of rbcS were present in the ancestral species of wheat. Thus, it is concluded that the divergence between the classes originated from an interchromosomal duplication and that several intrachromosomal duplications followed; these events occurred before the speciation of diploids. The detection of nonfunctional genes and the elimination of fragments suggests that several mechanisms were involved in reducing the copy number of rbcS during the evolution of wheat.

Base Sequence↗

Population genetics of multigene families that are dispersed into two or more chromosomes.

The evolution of multigene families whose members are dispersed into two or more nonhomologous chromosomes is studied from the standpoint of population genetics. By using a simple model of gene conversion, equilibrium and transient properties of the probability of identity of genes belonging to the family are investigated. Also, the time until fixation of a mutant belonging to a subdivided multigene family is studied by an approximation method. It is shown that dispersion of the family into two or more chromosomes has a relatively minor effect on the extent of identity between genes, unless the conversion rate between genes on nonhomologous chromosomes is extremely low, or unless the number of nonhomologous chromosomes on which gene members are dispersed is large.

Animals↗

Molecular characterization of rat multigene family encoding proline-rich proteins.

Three members of the rat proline-rich protein multigene family have been characterized. Each of these genes, RP4, RP13, and RP15, contains three exons and they are approximately 4.8, 5.7, and 5.4 kb, respectively. The DAN sequences of RP4 and RP13 are greater than 93% homologous in the 3.1-kb segment extending from the 5'-upstream region (approximately nucleotide -930) to 238 nucleotides after the second exon/intron junction; however, regions further downstream, intron II and exon III, share less than 43% identity. In contrast, exon III from RP15, RP13, and the previously sequenced mouse PRP gene MP2 are more than 73% conserved. These analyses suggest that the duplication of the ancestral genes to RP13 and RP4 occurred prior to the divergence of the rat PRP genes. The results also indicate that in the past 21.5 million years, multiple recombination events have resulted in a very high degree of divergence among intron II and exon III of RP4 and RP13. This divergence is due in part to the insertion of members of the rat long interspersed repeat DNA family at -930 bp upstream from the transcription initiation site and within intron II of RP13. Comparisons of the nucleotide sequences and organization of exon I with the genomic organization of PRP and glutamic acid/glutamine-rich protein genes in this and previous studies reveal striking resemblance among these genes. These observations are consistent with the notion that this super multigene family arose from duplication of progenitor genes via unequal crossing over events. In addition, the results suggest that concerted evolution has occurred within the tandemly repeated motif of exon II.

Amino Acid Sequence↗

Evolution of substrate recognition across a multigene family of glycosyltransferases in Arabidopsis.

The complete sequence of the Arabidopsis genome enables definitive characterization of multigene families and analysis of their phylogenetic relationships. Using a consensus sequence previously defined for glycosyltransferases that use small-molecular-weight acceptors, 107 gene sequences were identified in the Arabidopsis genome and used to construct a phylogenetic tree. Screening recombinant proteins for their catalytic activities in vitro has revealed enzymes active toward physiologically important substrates, including hormones and secondary metabolites. The aim of this study has been to use the phylogenetic relationships across the entire family to explore the evolution of substrate recognition and regioselectivity of glucosylation. Hydroxycoumarins have been used as the model substrates for the analysis in which 90 sequences have been assayed and 48 sequences shown to recognize these compounds. The study has revealed activity in 6 of the 14 phylogenetic groups of the multigene family, suggesting that basic features of substrate recognition are retained across substantial evolutionary periods.

Arabidopsis↗

The concerted evolution of 5S ribosomal genes linked to the repeat units of other multigene families.

We review all instances in which the nuclear 5S rRNA genes of fungi, protist, nematode, and arthropod species have been reported to be linked to the tandemly repeated units of the rDNA, trans-spliced leader, and histone multigene families. The evolution of these gene arrangements is analyzed by mapping them to independently derived phylogenies. These analyses show that 5S rRNA genes have repeatedly become linked to diverse tandemly repeated gene families and that such linkages have also been subsequently inverted or lost in some species. These variable gene linkages are probably the result of stochastic gains and losses of variant repeat units, where functional 5S rRNA had transposed, by the mechanisms which are responsible for the concerted evolution of tandemly repeated multigene families. We discuss the possible mechanisms of 5S rRNA gene transposition and suggest that the characteristics of their promoter elements, transcription, and termination signals may allow functional copies of these genes to be fortuitously transposed through an RNA intermediate. We also review the evidence which shows that the linked 5S rRNA gene copies are transcribed. We conclude that the observed patterns of 5S rRNA gene linkages to the repeat units of other tandemly repeated multigene families have likely arisen due to fortuitous recombination events and are unlikely to represent the remnants of an eubacterial-like arrangement of rDNA operons or to have been established due to selective pressures.

Biological Evolution↗

The PE multigene family: a 'molecular mantra' for mycobacteria.

The PE multigene family of Mycobacterium tuberculosis is remarkable in that it is composed of approximately 100 highly homologous genes that are found only in mycobacteria. Early evidence suggests that proteins encoded by certain members of this gene family could be present in the mycobacterial cell wall, impact antigen-presentation pathways and the ensuing host immune responses, and also provide a mechanism for generating antigenic diversity in mycobacteria.

Amino Acid Sequence↗