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Bioinformatic and phylogenetic analysis of the CLAVATA3/EMBRYO-SURROUNDING REGION (CLE) and the CLE-LIKE signal peptide genes in the Pinophyta.

Strabala TJ, Phillips L, West M, Stanbra L - BMC Plant Biol. (2014)

Bottom Line: The CLE and CLEL genes are found in conifers and they exhibit at least as much sequence diversity in these species as they do in other plant species.The preferential expression of these vascular development-regulating genes in phloem in conifers, as they are in dicot species, suggests close parallels in the regulation of secondary growth and wood formation in gymnosperm and dicot plants.Based on our bioinformatic analysis, we predict a novel mechanism of regulation of the expression of several conifer CLEL peptides, via alternative splicing resulting in the selection of alternative C-terminal exons encoding separate CLEL peptides.

View Article: PubMed Central - HTML - PubMed

Affiliation: Scion, 49 Sala St, PO Box 3020, Rotorua 3010, New Zealand. strabala@clear.net.nz.

ABSTRACT

Background: There is a rapidly growing awareness that plant peptide signalling molecules are numerous and varied and they are known to play fundamental roles in angiosperm plant growth and development. Two closely related peptide signalling molecule families are the CLAVATA3-EMBRYO-SURROUNDING REGION (CLE) and CLE-LIKE (CLEL) genes, which encode precursors of secreted peptide ligands that have roles in meristem maintenance and root gravitropism. Progress in peptide signalling molecule research in gymnosperms has lagged behind that of angiosperms. We therefore sought to identify CLE and CLEL genes in gymnosperms and conduct a comparative analysis of these gene families with angiosperms.

Results: We undertook a meta-analysis of the GenBank/EMBL/DDBJ gymnosperm EST database and the Picea abies and P. glauca genomes and identified 93 putative CLE genes and 11 CLEL genes among eight Pinophyta species, in the genera Cryptomeria, Pinus and Picea. The predicted conifer CLE and CLEL protein sequences had close phylogenetic relationships with their homologues in Arabidopsis. Notably, perfect conservation of the active CLE dodecapeptide in presumed orthologues of the Arabidopsis CLE41/44-TRACHEARY ELEMENT DIFFERENTIATION (TDIF) protein, an inhibitor of tracheary element (xylem) differentiation, was seen in all eight conifer species. We cloned the Pinus radiata CLE41/44-TDIF orthologues. These genes were preferentially expressed in phloem in planta as expected, but unexpectedly, also in differentiating tracheary element (TE) cultures. Surprisingly, transcript abundances of these TE differentiation-inhibitors sharply increased during early TE differentiation, suggesting that some cells differentiate into phloem cells in addition to TEs in these cultures. Applied CLE13 and CLE41/44 peptides inhibited root elongation in Pinus radiata seedlings. We show evidence that two CLEL genes are alternatively spliced via 3'-terminal acceptor exons encoding separate CLEL peptides.

Conclusions: The CLE and CLEL genes are found in conifers and they exhibit at least as much sequence diversity in these species as they do in other plant species. Only one CLE peptide sequence has been 100% conserved between gymnosperms and angiosperms over 300 million years of evolutionary history, the CLE41/44-TDIF peptide and its likely conifer orthologues. The preferential expression of these vascular development-regulating genes in phloem in conifers, as they are in dicot species, suggests close parallels in the regulation of secondary growth and wood formation in gymnosperm and dicot plants. Based on our bioinformatic analysis, we predict a novel mechanism of regulation of the expression of several conifer CLEL peptides, via alternative splicing resulting in the selection of alternative C-terminal exons encoding separate CLEL peptides.

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Phylogenetic analysis of Arabidopsis and Pinophyta CLE and CLEL proteins. A 1000-iteration Neighbour-Joining analysis using the Poisson correction method with alignment gaps and missing data eliminated only in pairwise sequence comparisons was used to create bootstrap consensus trees representing the putative phylogenetic relationships among the CLE and CLEL proteins between Arabidopsis thaliana and the Pinophyta species. The trees are drawn to scale, with branch lengths in the units of the number of amino acid substitutions per site. Arabidopsis proteins are represented by black squares and Pinophyta proteins are represented by red squares. A. CLE proteins; 194 positions in the final dataset. B. CLEL proteins; 277 positions in the final dataset.
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Figure 4: Phylogenetic analysis of Arabidopsis and Pinophyta CLE and CLEL proteins. A 1000-iteration Neighbour-Joining analysis using the Poisson correction method with alignment gaps and missing data eliminated only in pairwise sequence comparisons was used to create bootstrap consensus trees representing the putative phylogenetic relationships among the CLE and CLEL proteins between Arabidopsis thaliana and the Pinophyta species. The trees are drawn to scale, with branch lengths in the units of the number of amino acid substitutions per site. Arabidopsis proteins are represented by black squares and Pinophyta proteins are represented by red squares. A. CLE proteins; 194 positions in the final dataset. B. CLEL proteins; 277 positions in the final dataset.

Mentions: We examined the phylogenetic relationships to Arabidopsis among the conifer CLE and CLEL precursor protein sequences as a first attempt to assess potential protein role(s) and/or function(s). A 1000-iteration Neighbour-Joining analysis grouped the conifer protein sequences with varying degrees of phylogenetic distance from the Arabidopsis CLE and CLEL clades (Figure 4). In particular, a large clade of 30 protein sequences was grouped with Arabidopsis CLE41 and CLE42 proteins and 39 other conifer proteins were grouped with Arabidopsis CLE20 (Figures 4A and5). Among the CLEL proteins, the closest Arabidopsis - Pinophyta evolutionary relationship was seen with P. glauca CLEL16 and Arabidopsis RGF4 (Figure 4B). CLEL17 grouped with CLE18, but this relationship may be spurious, as CLEL17 is only a partial protein sequence.


Bioinformatic and phylogenetic analysis of the CLAVATA3/EMBRYO-SURROUNDING REGION (CLE) and the CLE-LIKE signal peptide genes in the Pinophyta.

Strabala TJ, Phillips L, West M, Stanbra L - BMC Plant Biol. (2014)

Phylogenetic analysis of Arabidopsis and Pinophyta CLE and CLEL proteins. A 1000-iteration Neighbour-Joining analysis using the Poisson correction method with alignment gaps and missing data eliminated only in pairwise sequence comparisons was used to create bootstrap consensus trees representing the putative phylogenetic relationships among the CLE and CLEL proteins between Arabidopsis thaliana and the Pinophyta species. The trees are drawn to scale, with branch lengths in the units of the number of amino acid substitutions per site. Arabidopsis proteins are represented by black squares and Pinophyta proteins are represented by red squares. A. CLE proteins; 194 positions in the final dataset. B. CLEL proteins; 277 positions in the final dataset.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4016512&req=5

Figure 4: Phylogenetic analysis of Arabidopsis and Pinophyta CLE and CLEL proteins. A 1000-iteration Neighbour-Joining analysis using the Poisson correction method with alignment gaps and missing data eliminated only in pairwise sequence comparisons was used to create bootstrap consensus trees representing the putative phylogenetic relationships among the CLE and CLEL proteins between Arabidopsis thaliana and the Pinophyta species. The trees are drawn to scale, with branch lengths in the units of the number of amino acid substitutions per site. Arabidopsis proteins are represented by black squares and Pinophyta proteins are represented by red squares. A. CLE proteins; 194 positions in the final dataset. B. CLEL proteins; 277 positions in the final dataset.
Mentions: We examined the phylogenetic relationships to Arabidopsis among the conifer CLE and CLEL precursor protein sequences as a first attempt to assess potential protein role(s) and/or function(s). A 1000-iteration Neighbour-Joining analysis grouped the conifer protein sequences with varying degrees of phylogenetic distance from the Arabidopsis CLE and CLEL clades (Figure 4). In particular, a large clade of 30 protein sequences was grouped with Arabidopsis CLE41 and CLE42 proteins and 39 other conifer proteins were grouped with Arabidopsis CLE20 (Figures 4A and5). Among the CLEL proteins, the closest Arabidopsis - Pinophyta evolutionary relationship was seen with P. glauca CLEL16 and Arabidopsis RGF4 (Figure 4B). CLEL17 grouped with CLE18, but this relationship may be spurious, as CLEL17 is only a partial protein sequence.

Bottom Line: The CLE and CLEL genes are found in conifers and they exhibit at least as much sequence diversity in these species as they do in other plant species.The preferential expression of these vascular development-regulating genes in phloem in conifers, as they are in dicot species, suggests close parallels in the regulation of secondary growth and wood formation in gymnosperm and dicot plants.Based on our bioinformatic analysis, we predict a novel mechanism of regulation of the expression of several conifer CLEL peptides, via alternative splicing resulting in the selection of alternative C-terminal exons encoding separate CLEL peptides.

View Article: PubMed Central - HTML - PubMed

Affiliation: Scion, 49 Sala St, PO Box 3020, Rotorua 3010, New Zealand. strabala@clear.net.nz.

ABSTRACT

Background: There is a rapidly growing awareness that plant peptide signalling molecules are numerous and varied and they are known to play fundamental roles in angiosperm plant growth and development. Two closely related peptide signalling molecule families are the CLAVATA3-EMBRYO-SURROUNDING REGION (CLE) and CLE-LIKE (CLEL) genes, which encode precursors of secreted peptide ligands that have roles in meristem maintenance and root gravitropism. Progress in peptide signalling molecule research in gymnosperms has lagged behind that of angiosperms. We therefore sought to identify CLE and CLEL genes in gymnosperms and conduct a comparative analysis of these gene families with angiosperms.

Results: We undertook a meta-analysis of the GenBank/EMBL/DDBJ gymnosperm EST database and the Picea abies and P. glauca genomes and identified 93 putative CLE genes and 11 CLEL genes among eight Pinophyta species, in the genera Cryptomeria, Pinus and Picea. The predicted conifer CLE and CLEL protein sequences had close phylogenetic relationships with their homologues in Arabidopsis. Notably, perfect conservation of the active CLE dodecapeptide in presumed orthologues of the Arabidopsis CLE41/44-TRACHEARY ELEMENT DIFFERENTIATION (TDIF) protein, an inhibitor of tracheary element (xylem) differentiation, was seen in all eight conifer species. We cloned the Pinus radiata CLE41/44-TDIF orthologues. These genes were preferentially expressed in phloem in planta as expected, but unexpectedly, also in differentiating tracheary element (TE) cultures. Surprisingly, transcript abundances of these TE differentiation-inhibitors sharply increased during early TE differentiation, suggesting that some cells differentiate into phloem cells in addition to TEs in these cultures. Applied CLE13 and CLE41/44 peptides inhibited root elongation in Pinus radiata seedlings. We show evidence that two CLEL genes are alternatively spliced via 3'-terminal acceptor exons encoding separate CLEL peptides.

Conclusions: The CLE and CLEL genes are found in conifers and they exhibit at least as much sequence diversity in these species as they do in other plant species. Only one CLE peptide sequence has been 100% conserved between gymnosperms and angiosperms over 300 million years of evolutionary history, the CLE41/44-TDIF peptide and its likely conifer orthologues. The preferential expression of these vascular development-regulating genes in phloem in conifers, as they are in dicot species, suggests close parallels in the regulation of secondary growth and wood formation in gymnosperm and dicot plants. Based on our bioinformatic analysis, we predict a novel mechanism of regulation of the expression of several conifer CLEL peptides, via alternative splicing resulting in the selection of alternative C-terminal exons encoding separate CLEL peptides.

Show MeSH