Genome-wide identification of key genes related to chloride ion (Cl-) channels and transporters in response to salt stress in birch

Abstract

Background: Soil salinization is a common matter of concern all over the world, which severely affects plant production, soil health, and ecosystem stability. Birch is a significant afforestation tree species with great ecological, economic, and evolutionary value. It is an excellent model system for studying the acclimation and adaptation of woody plants to extreme environmental conditions due to the current advancements in genomics, genetic variability, and extensive studies with a focus on various biotic and abiotic stresses. To date, the genetic regulation of birch trees in defending against environmental stimuli, particularly salt stress, has made great progress. However, the information on how genes related to chloride ion (Cl^-) channels and transporters respond to salt stress remains poorly understood in birch.

Results: Herein, we performed the genome-wide identification, evolutionary relationship, sequence analysis, and expression patterns of seven family genes encoding Cl^- channels or transporters in birch. We identified one, one, four, seven, 13, 56, and 67 genes belonging to the cation chloride co-transporter (CCC) family, bestrophin family, slow type anion channel (SLAC) family, chloride channel (CLC) family, aluminum-activated malate transporter (ALMT) family, nitrate transporter 1/peptide transporter family (NPF) family, and multidrug and toxic compound extrusion (MATE ) family, respectively. Except for motif prediction, a high similarity of the conserved domain composition, gene structures, and physicochemical properties was observed within each group of a certain gene family. Gene Ontology (GO) classification analysis showed that the genes encoding ALMTs, CCCs, CLCs, MATEs, NPFs, and SLACs were mainly involved in ion transport. Transcriptome profiles showed that there were 52 genes related to Cl^- transporters and channels differentially expressed in any tissue of the two birch species subjected to salt treatment. Of these, three genes, designated as BpCLCa/b, BpDTX45, and BpDTX18/19, were simultaneously expressed at altered levels in all tissues of the two birch species, which was consistent with the results of quantitative real-time polymerase chain reaction (RT-qPCR) assay. In addition, the molecular docking simulations showed that sodium ion (Na⁺) and Cl^-, respectively, had a strong binding affinity towards BpCLCa/b, BpDTX45, and BpDTX18/19.

Conclusion: A total of 149 genes related to Cl^- channels and transporters were identified from the birch genome. We proposed BpCLCa/b, BpDTX45, and BpDTX18/19 as ion channels or transporters in controlling ionic homeostasis during salt stress response. The findings of our study can provide valuable resources for genetic improvement of salt-tolerant birch varieties for high-quality plantation forests in harsh environments.

Keywords: Birch; Chlorine; Ion channel; Salt stress; Sodium; Transporter.

Fig. 1

Identification and analysis of ALMT family in birch. a Phylogenetic tree of ALMT family. Two proteins of 126,297 and 56,569 were used as an outgroup. Black solid star, square, and circle represented the proteins from Arabidopsis, poplar, and birch, respectively. b Gene structures of BpALMTs. c Conserved motifs of BpALMTs. Each motif was represented by a number in colored box

Fig. 2

Identification and analysis of SLAC, CCC, and bestrophin family in birch. a Phylogenetic tree of SLAC family. Two proteins of WP_165280682.1 and WP_090253539.1 were used as an outgroup. b Phylogenetic tree of CCC family. Three proteins of evm 27.model.AmTr v1.0 scaffold00092.18, AmTrH1.02G015400.1.p, and Mapoly0050s0083.1.p were used as an outgroup. c Phylogenetic tree of bestrophin family. Two proteins of evm 27.model.AmTr v1.0 scaffold00009.260 and AmTrH1.07G115300.1 were used as an outgroup. d Gene structures of BpSLACs. e Conserved motifs of BpSLACs. f Gene structures of the BpCCC. g Conserved motifs of the BpCCC. h Gene structures of the BpBest. i Conserved motifs of the BpBest. Black solid star, square, and circle represented the proteins from Arabidopsis, poplar, and birch, respectively. Each conserved motif was represented by a colored box

Fig. 3

Identification and analysis of CLC family in birch. a Phylogenetic tree of CLC family. The protein sequence of Bpev01.c0726.g0012.g0013 was revised by Bpev01.c0726.g0012 and Bpev01.c0726.g0013. Two proteins of WP_066121478.1 and WP_308254669.1 were used as an outgroup. Black solid star, square, and circle represented the proteins from Arabidopsis, poplar, and birch, respectively. b Gene structures of BpCLCs. c Conserved motifs of BpCLCs. Each motif was represented by a colored box

Fig. 4

Phylogenetic analysis of MATE family in Arabidopsis, birch, and poplar. The protein sequence of Bpev01.c2011.g0004.g0003 was revised by Bpev01.c2011.g0004 and Bpev01.c2011.g0003. The protein sequence of Bpev01.c0630.g0013.g0014.g0015 was revised by Bpev01.c0630.g0013, Bpev01.c0630.g0014, and Bpev01.c0630.g0015. The protein sequence of Bpev01.c0302.g0001.g0002 was revised by Bpev01.c0302.g0001 and Bpev01.c0302.g0002. The protein sequence of Bpev01.c1385.g0003.g0002 was revised by Bpev01.c1385.g0003 and Bpev01.c1385.g0002. Black solid star, square, and circle represented the proteins from Arabidopsis, poplar, and birch, respectively

Fig. 5

Gene structure and motif analysis of MATE family in birch. a Gene structures of BpMATEs. b Conserved motifs of BpMATEs. Each motif was represented by a colored box. Gene ID in green, blue, purple, and orange color represented the group I, II, III, and IV, respecitvely

Fig. 6

Phylogenetic analysis of NPF family in Arabidopsis, birch, and poplar. The protein sequence of Bpev01.c0389.g0061.g0060 was revised by Bpev01.c0389.g0061 and Bpev01.c0389.g0060. Black solid star, square, and circle represented the proteins from Arabidopsis, poplar, and birch, respectively

Fig. 7

Gene structure and motif analysis of NPF family in birch. a Gene structures of BpNPFs. b Conserved motifs of BpNPFs. Each motif was represented by a colored box. Gene ID in green, blue, purple, and orange color represented the group I, II, III, and IV, respecitvely

Fig. 8

Identification of candidate genes related to Cl⁻ channels and transporters and molecular docking simulation of their proteins in birch. a Expression patterns of genes related to Cl⁻ channels and transporters in birch after salt treatment. Bpev01.c2011.g0004_partial represented Bpev01.c2011.g0004, which was a part of Bpev01.c2011.g0004.g0003. The expression values were indicated by log₂|fold change| values. b Relative expression levels of candidate genes in roots and leaves of B. platyphylla after salt stress. c The molecular docking simulation. The dashed line represented the coordinate covalent bonds