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The homology-based analysis identifies 41 putative AQPs in the garden pea genome. Among them, all but two genes (Psat0s3550g0040.1, Psat0s2987g0040.1) encode full-length aquaporin-like sequences (Table 1). The conserved protein domain analysis later validated all of the expected AQPs (Supplemental Table S2). To systematically classify these genes and elucidate their relationship with the AQPs from other plants' a phylogenetic tree was created. It clearly showed that the AQPs from pea and its close relative M. truncatula formed four distinct clusters, which represented the different subfamilies of AQPs i.e. TIPs, PIPs, NIPs, and SIPs (Fig. 1a). However, out of the 41 identified pea AQPs, 4 AQPs couldn't be tightly aligned with the Medicago AQPs and thus were put to a new phylogenetic tree constructed with AQPs from rice, Arabidopsis, and soybean. This additional analysis assigned one of the 4 AQPs to the XIP subfamily and the rest three to the TIP or NIP subfamilies (Fig. 1b). Therefore, it is concluded that the 41 PsAQPs comprise 11 PsTIPs, 15 PsNIPs, 9 PsPIPs, 5 PsSIPs, and 1 PsXIP (Table 2). The PsPIPs formed two major subgroups namely PIP1s and PIP2s, which comprise three and six members, respectively (Table 1). The PsTIPs formed two major subgroups TIPs 1 (PsTIP1-1, PsTIP1-3, PsTIP1-4, PsTIP1-7) and TIPs 2 (PsTIP2-1, PsTIP2-2, PsTIP2-3, PsTIP2-6) each having four members (Table 2). Detailed information such as gene/protein names, accession numbers, the length of deduced polypeptides, and protein structural features are presented in Tables 1 & 2
Table 1. Description and distribution of aquaporin genes identified in the garden pea genome.
Chromosome S. No Gene Name Gene ID Gene length
(bp)Location Start End Transcription length (bp) CDS length
(bp)Protein length
(aa)1 PsPIP1-1 Psat5g128840.3 2507 chr5LG3 231,127,859 231,130,365 675 675 225 2 PsPIP1-2 Psat2g034560.1 1963 chr2LG1 49,355,958 49,357,920 870 870 290 3 PsPIP1-4 Psat2g182480.1 1211 chr2LG1 421,647,518 421,648,728 864 864 288 4 PsPIP2-1 Psat6g183960.1 3314 chr6LG2 369,699,084 369,702,397 864 864 288 5 PsPIP2-2-1 Psat4g051960.1 1223 chr4LG4 86,037,446 86,038,668 585 585 195 6 PsPIP2-2-2 Psat5g279360.2 2556 chr5LG3 543,477,849 543,480,404 2555 789 263 7 PsPIP2-3 Psat7g228600.2 2331 chr7LG7 458,647,213 458,649,543 2330 672 224 8 PsPIP2-4 Psat3g045080.1 1786 chr3LG5 100,017,377 100,019,162 864 864 288 9 PsPIP2-5 Psat0s3550g0040.1 1709 scaffold03550 20,929 22,637 1191 1191 397 10 PsTIP1-1 Psat3g040640.1 2021 chr3LG5 89,426,473 89,428,493 753 753 251 11 PsTIP1-3 Psat3g184440.1 2003 chr3LG5 393,920,756 393,922,758 759 759 253 12 PsTIP1-4 Psat7g219600.1 2083 chr7LG7 441,691,937 441,694,019 759 759 253 13 PsTIP1-7 Psat6g236600.1 1880 chr6LG2 471,659,417 471,661,296 762 762 254 14 PsTIP2-1 Psat1g005320.1 1598 chr1LG6 7,864,810 7,866,407 750 750 250 15 PsTIP2-2 Psat4g198360.1 1868 chr4LG4 407,970,525 407,972,392 750 750 250 16 PsTIP2-3 Psat1g118120.1 2665 chr1LG6 230,725,833 230,728,497 768 768 256 17 PsTIP2-6 Psat2g177040.1 1658 chr2LG1 416,640,482 416,642,139 750 750 250 18 PsTIP3-2 Psat6g054400.1 1332 chr6LG2 54,878,003 54,879,334 780 780 260 19 PsTIP4-1 Psat6g037720.2 1689 chr6LG2 30,753,624 30,755,312 1688 624 208 20 PsTIP5-1 Psat7g157600.1 1695 chr7LG7 299,716,873 299,718,567 762 762 254 21 PsNIP1-1 Psat1g195040.2 1864 chr1LG6 346,593,853 346,595,716 1863 645 215 22 PsNIP1-3 Psat1g195800.1 1200 chr1LG6 347,120,121 347,121,335 819 819 273 23 PsNIP1-5 Psat7g067480.1 2365 chr7LG7 109,420,633 109,422,997 828 828 276 24 PsNIP1-6 Psat7g067360.1 2250 chr7LG7 109,270,462 109,272,711 813 813 271 25 PsNIP1-7 Psat1g193240.1 1452 chr1LG6 344,622,606 344,624,057 831 831 277 26 PsNIP2-1-2 Psat3g197520.1 669 chr3LG5 420,092,382 420,093,050 345 345 115 27 PsNIP2-2-2 Psat3g197560.1 716 chr3LG5 420,103,168 420,103,883 486 486 162 28 PsNIP3-1 Psat2g072000.1 1414 chr2LG1 133,902,470 133,903,883 798 798 266 29 PsNIP4-1 Psat7g126440.1 1849 chr7LG7 209,087,362 209,089,210 828 828 276 30 PsNIP4-2 Psat5g230920.1 1436 chr5LG3 463,340,575 463,342,010 825 825 275 31 PsNIP5-1 Psat6g190560.1 1563 chr6LG2 383,057,323 383,058,885 867 867 289 32 PsNIP6-1 Psat5g304760.4 5093 chr5LG3 573,714,868 573,719,960 5092 486 162 33 PsNIP6-2 Psat7g036680.1 2186 chr7LG7 61,445,341 61,447,134 762 762 254 34 PsNIP6-3 Psat7g259640.1 2339 chr7LG7 488,047,315 488,049,653 918 918 306 35 PsNIP7-1 Psat6g134160.2 4050 chr6LG2 260,615,019 260,619,068 4049 1509 503 36 PsSIP1-1 Psat3g091120.1 3513 chr3LG5 187,012,329 187,015,841 738 738 246 37 PsSIP1-2 Psat1g096840.1 3609 chr1LG6 167,126,599 167,130,207 744 744 248 38 PsSIP1-3 Psat7g203280.1 2069 chr7LG7 401,302,247 401,304,315 720 720 240 39 PsSIP2-1-1 Psat0s2987g0040.1 706 scaffold02987 177,538 178,243 621 621 207 40 PsSIP2-1-2 Psat3g082760.1 3135 chr3LG5 173,720,100 173,723,234 720 720 240 41 PsXIP2-1 Psat7g178080.1 2077 chr7LG7 335,167,251 335,169,327 942 942 314 bp: base pair, aa: amino acid. Figure 1.
Phylogenetic analysis of the identified AQPs from pea genome. (a) The pea AQPs proteins aligned with those from the cool-season legume Medicago truncatual. (b) The four un-assigned pea AQPs in (a) (denoted as NA) were further aligned with the AQPs of rice, soybean, and Arabidopsis by using the Clustal W program implemented in MEGA 7 software. The nomenclature of PsAQPs was based on homology with the identified aquaporins that were clustered together.
Table 2. Protein information, conserved amino acid residues, trans-membrane domains, selectivity filter, and predicted subcellular localization of the 39 full-length pea aquaporins.
S. No AQPs Gene Length TMH NPA NPA ar/R selectivity filter pI WoLF PSORT Plant-mPLoc LB LE H2 H5 LE1 LE2 Plasma membrane intrinsic proteins (PIPs) 1 PsPIP1-1 Psat5g128840.3 225 4 NPA 0 F 0 0 0 8.11 Plas Plas 2 PsPIP1-2 Psat2g034560.1 290 2 NPA NPA F H T R 9.31 Plas Plas 3 PsPIP1-4 Psat2g182480.1 288 6 NPA NPA F H T R 9.29 Plas Plas 4 PsPIP2-1 Psat6g183960.1 288 6 NPA NPA F H T 0 8.74 Plas Plas 5 PsPIP2-2-1 Psat4g051960.1 195 3 0 0 F H T R 8.88 Plas Plas 6 PsPIP2-2-2 Psat5g279360.2 263 5 NPA NPA F H T R 5.71 Plas Plas 7 PsPIP2-3 Psat7g228600.2 224 4 NPA 0 F F 0 0 6.92 Plas Plas 8 PsPIP2-4 Psat3g045080.1 288 6 NPA NPA F H T R 8.29 Plas Plas Tonoplast intrinsic proteins (TIPs) 1 PsTIP1-1 Psat3g040640.1 251 7 NPA NPA H I A V 6.34 Plas Vacu 2 PsTIP1-3 Psat3g184440.1 253 6 NPA NPA H I A V 5.02 Plas/Vacu Vacu 3 PsTIP1-4 Psat7g219600.1 253 7 NPA NPA H I A V 4.72 Vacu Vacu 4 PsTIP1-7 Psat6g236600.1 254 6 NPA NPA H I A V 5.48 Plas/Vacu Vacu 5 PsTIP2-1 Psat1g005320.1 250 6 NPA NPA H I G R 8.08 Vacu Vacu 6 PsTIP2-2 Psat4g198360.1 250 6 NPA NPA H I G R 5.94 Plas/Vacu Vacu 7 PsTIP2-3 Psat1g118120.1 256 6 NPA NPA H I A L 6.86 Plas/Vacu Vacu 8 PsTIP2-6 Psat2g177040.1 250 6 NPA NPA H I G R 4.93 Vacu Vacu 9 PsTIP3-2 Psat6g054400.1 260 6 NPA NPA H I A R 7.27 Plas/Vacu Vacu 10 PsTIP4-1 Psat6g037720.2 208 6 NPA NPA H I A R 6.29 Vac/ plas Vacu 11 PsTIP5-1 Psat7g157600.1 254 7 NPA NPA N V G C 8.2 Vacu /plas Vacu/Plas Nodulin-26 like intrisic proteins (NIPs) 1 PsNIP1-1 Psat1g195040.2 215 5 NPA 0 W V F 0 6.71 Plas Plas 2 PsNIP1-3 Psat1g195800.1 273 5 NPA NPV W V A R 6.77 Plas Plas 3 PsNIP1-5 Psat7g067480.1 276 6 NPA NPV W V A N 8.98 Plas Plas 4 PsNIP1-6 Psat7g067360.1 271 6 NPA NPA W V A R 8.65 Plas/Vacu Plas 5 PsNIP1-7 Psat1g193240.1 277 6 NPA NPA W I A R 6.5 Plas/Vacu Plas 6 PsNIP2-1-2 Psat3g197520.1 115 2 NPA O G 0 0 0 9.64 Plas Plas 7 PsNIP2-2-2 Psat3g197560.1 162 3 0 NPA 0 S G R 6.51 Plas Plas 8 PsNIP3-1 Psat2g072000.1 266 5 NPA NPA S I A R 8.59 Plas/Vacu Plas 9 PsNIP4-1 Psat7g126440.1 276 6 NPA NPA W V A R 6.67 Plas Plas 10 PsNIP4-2 Psat5g230920.1 275 6 NPA NPA W L A R 7.01 Plas Plas 11 PsNIP5-1 Psat6g190560.1 289 5 NPS NPV A I G R 7.1 Plas Plas 12 PsNIP6-1 Psat5g304760.4 162 2 NPA 0 I 0 0 0 9.03 Plas Plas 13 PsNIP6-2 Psat7g036680.1 254 0 0 0 G 0 0 0 5.27 Chlo Plas/Nucl 14 PsNIP6-3 Psat7g259640.1 306 6 NPA NPV T I G R 8.32 Plas Plas 15 PsNIP7-1 Psat6g134160.2 503 0 NLK 0 W G Q R 8.5 Vacu Chlo/Nucl Small basic intrinsic proteins (SIPs) 1 PsSIP1-1 Psat3g091120.1 246 6 NPT NPA V L P N 9.54 Plas Plas/Vacu 2 PsSIP1-2 Psat1g096840.1 248 5 NTP NPA I V P L 9.24 Vacu Plas/Vacu 3 PsSIP1-3 Psat7g203280.1 240 6 NPS NPA N L P N 10.32 Chlo Plas 4 PsSIP2-1-2 Psat3g082760.1 240 4 NPL NPA Y L G S 10.28 Plas Plas Uncharacterized X intrinsic proteins (XIPs) 1 PsXIP2-1 Psat7g178080.1 314 6 SPV NPA V V R M 7.89 Plas Plas Length: protein length (aa); pI: Isoelectric point; Trans-membrane helicase (TMH) represents for the numbers of Trans-membrane helices predicted by TMHMM Server v.2.0 tool; WoLF PSORT and Plant-mPLoc: best possible cellualr localization predicted by the WoLF PSORT and Plant-mPLoc tool, respectively (Chlo Chloroplast, Plas Plasma membrane, Vacu Vacuolar membrane, Nucl Nucleus); LB: Loop B, L: Loop E; NPA: Asparagine-Proline-Alanine; H2 represents for Helix 2, H5 represents for Helix 5, LE1 represents for Loop E1, LE2 represents for Loop E2, Ar/R represents for Aromatic/Arginine. Genome distribution and gene structure analysis of pea AQPs
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To understand the genome distribution of the 41 PsAQPs, we mapped these genes onto the seven chromosomes of a pea to retrieve their physical locations (Fig. 2). The greatest number (10) of AQPs were found on chromosome 7, whereas the least (2) on chromosome 4 (Fig. 2 and Table 1). Chromosomes 1 and 6 each contain six aquaporin genes, whereas chromosomes 2, 3, and 5 carry four, seven, and four aquaporin genes, respectively (Fig. 2). The trend of clustered distribution of AQPs was seen on specific chromosomes, particularly near the end of chromosome 7.
Figure 2.
Chromosomal localization of the 41 PsAQPs on the seven chromosomes of pea. Chr1-7 represents the chromosomes 1 to 7. The numbers on the right of each chromosome show the physical map positions of the AQP genes (Mbp). Blue, green, orange, brown, and black colors represent TIPs, NIPs, PIPs, SIPs, and XIP, respectively.
The 39 full-length PsAQP proteins have a length of amino acid ranging from 115 to 503 (Table 1) and Isoelectric point (pI) values ranging from 4.72 to 10.35 (Table 2). As a structural signature, transmembrane domains were predicted to exist in all PsAQPs, with the number in individual AQPs varying from 2 to 6. By subfamilies, TIPs harbor the greatest number of TM domains in total, followed by PIPs, NIPs, SIPs, and XIP (Table 2). Exon-intron structure analysis showed that most PsAQPs (16/39) having two introns, while ten members had three, seven members had four, and five members had only one intron (Fig. 3). Overall, PsAQPs exhibited a complex structure with varying intron numbers, positions, and lengths.
Figure 3.
The exon-intron structures of the AQP genes in pea. Upstream/downstream region, exon, and intron are represented by a blue box, yellow box, and grey line, respectively.
Characterization of the NPA motifs
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As aforementioned, generally highly conserved two NPA motifs generate an electrostatic repulsion of protons in AQPs to form the water channel, which is essential for the transport of substrate molecules[15]. In order to comprehend the potential physiological function and substrate specificity of pea aquaporins, NPA motifs (LB, LE) and residues at the ar/R selectivity filter (H2, H5, LE1, and LE2) were examined. (Table 2). We found that all PsTIPs and most PsPIPs had two conserved NPA motifs except for PsPIP1-1, PsPIP2-2-1, and PsPIP2-3, each having a single NPA motif. Among PsNIPs, PsNIP1-6, PsNIP1-6, PsNIP1-7, PsNIP3-1, PsNIP4-1 and PSNIP4-2 had two NPA domains, while PsNIP1-1, PsNIP2-1-2, PsNIP2-2-2 and PsNIP6-1 each had a single NPA motif. In the PsNIP sub-family, the first NPA motif showed an Alanine (A) to Valine (V) substitution in three PsNIPs (PsNIP1-3, PsNIP1-5, and PsNIP6-3) (Table 2). Furthermore, the NPA domains of all members of the XIP and SIP subfamilies were different. The second NPA motif was conserved in PsSIP aquaporins, however, all of the first NPA motifs had Alanine (A) replaced by Leucine (L) (PsSIP2-1-1, PsSIP2-1-2) or Threonine (T) (PsSIP1-1). In comparison to other subfamilies, this motif variation distinguishes water and solute-transporting aquaporins[45].
Compared to NPA motifs, the ar/R positions were more variable and the amino acid composition appeared to be subfamily-dependent. The majority of PsPIPs had phenylalanine at H2, histidine at H5, threonine at LE1, and arginine at LE2 selective filter (Table 2). All of the PsTIP1 members had a Histidine-Isoleucine-Alanine-Valine structure at this position, while all PsTIP2 members but PsTIP2-3 harbored Histidine-Isoleucine-Glycine-Arginine. Similarly, PsNIPs, PsSIPs and PsXIP also showed subgroup-specific variation in ar/R selectivity filter (Table 2). Each of these substitutions partly determines the function of transporting water[46].
Predicted subcellular localization of PsAQPs
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Sequence-based subcellular localization analysis using WoLF PSORT predicted that all PsPIPs localized in the plasma membrane, which is consistent with their subfamily classification (Table 2). Around half (5/11) of the PsTIPs (PsTIP1-4, PsTIP2-1, PsTIP2-6, PsTIP4-1, and PsTIP5-1) were predicted to localize within vacuoles. However, several TIP members (PsTIP1-1, PsTIP1-3, PsTIP1-7, PsTIP2-2, PsTIP2-3 and PsTIP3-2) were predicted to localize in plasma membranes. We then further investigated their localizations by using another software (Plant-mPLoc, Table 2), which predicted that all the PsTIPs localize within vacuoles, thus supporting that they are tonoplast related. An overwhelming majority of PsNIPs (14/15) and PsXIP were predicted to be found only in plasma membranes., which was also expected (Table 2). Collectively, the versatility in subcellular localization of the pea AQPs is implicative of their distinct roles in controlling water and/or solute transport in the context of plant cell compartmentation.
Tissue expression profiles of PsAQPs
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Tissue expression patterns of genes are indicative of their functions. Since there were rich resources of RNA-Seq data from various types of pea tissues in the public database, they were used for the extraction of expression information of PsAQP genes as represented by FPKM values. A heat map was generated to show the expression patterns of PsAQP genes in 18 different tissues/stages and their responses to nitrate levels (Fig. 4). According to the heat map, PsPIP1-2, PsPIP2-3 were highly expressed in root and nodule G (Low-nitrate), whereas PsTIP1-4, PsTIP2-6, and PsNIP1-7 were only expressed in roots in comparison to other tissues. The result also demonstrated that PsPIP1-1 and PsNIP3-1 expressed more abundantly in leaf, tendril, and peduncle, whereas PsPIP2-2-2 and PsTIP1-1 showed high to moderate expressions in all the samples except for a few. Interestingly, PsTIP1-1 expression in many green tissues seemed to be oppressed by low-nitrate. In contrast, some AQPs such as PsTIP1-3, PsTIP1-7, PsTIP5-1, PsNIP1-5, PsNIP4-1, PsNIP5-1, and PsSIP2-1-1 showed higher expression only in the flower tissue. There were interesting developmental stage-dependent regulations of some AQPs in seeds (Fig. 4). For example, PsPIP2-1, PsPIP2-2-1, PsNIP1-6, PsSIP1-1, and PsSIP1-2 were more abundantly expressed in the Seed_12 dap (days after pollination;) tissue than in the Seed_5 dai (days after imbibition) tissue; reversely, PsPIP2-2-2, PsPIP2-4, PsTIP2-3, and PsTIP3-2 showed higher expression in seed_5 dai in compare to seed_12 dap tissues (Fig. 4). The AQP genes may have particular functional roles in the growth and development of the pea based on their tissue-specific expression.
Figure 4.
Heatmap analysis of the expression of pea AQP gene expressions in different tissues using RNA-seq data (PRJNA267198). Normalized expression of aquaporins in terms of reads per kilobase of transcript per million mapped reads (RPKM) showing higher levels of PIPs, NIPs, TIPs SIPs, and XIP expression across the different tissues analyzed. (Stage A represents 7-8 nodes; stage B represents the start of flowering; stage D represents germination, 5 d after imbibition; stage E represents 12 d after pollination; stage F represents 8 d after sowing; stage G represents 18 d after sowing, LN: Low-nitrate; HN: High-nitrate.
PsAQPs expressions in response to osmotic stress and fullerol treatment in imbibing embryos
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Expressions of plant AQPs in vegetative tissues under normal and stressed conditions have been extensively studied[15]; however, little is known about the transcriptional regulation of AQP genes in seeds/embryos. To provide insights into this specific area, wet-bench RNA-Seq was performed on the germinating embryo samples isolated from water (W)-imbibed seeds and those treated with mannitol (M, an osmotic reagent), mannitol, and mannitol plus fullerol (F, a nano-antioxidant). The phenotypic evaluation showed that M treatment had a substantial inhibitory effect on radicle growth, whereas the supplement of F significantly mitigated this inhibition at all concentrations, in particular, 100 mg/mL in MF3, which increased the radicle length by ~33% as compared to that under solely M treatment (Fig. 5). The expression values of PsAQP genes were removed from the RNA-Seq data, and pairwise comparisons were made within the Group 1: W vs M, and Group 2: W vs MF3, where a total of ten and nince AQPs were identified as differentially expressed genes (DEGs), respectively (Fig. 6). In Group 1, six DEGs were up-regulated and four DEGs down-regulated, whereas in Group 2, six DEGs were up-regulated and three DEGs down-regulated. Four genes viz. PsPIPs2-5, PsNIP6-3, PsTIP2-3, and PsTIP3-2 were found to be similarly regulated by M or MF3 treatment (Fig. 6), indicating that their regulation by osmotic stress couldn't be mitigated by fullerol. Three genes, all being PsNIPs (1-1, 2-1-2, and 4-2), were up-regulated only under mannitol treatment without fullerol, suggesting that their perturbations by osmotic stress were migrated by the antioxidant activities. In contrast, four other genes namely PsTIP2-2, PsTIP4-1, PsNIP1-5, and PsSIP1-3 were only regulated under mannitol treatment when fullerol was present.
Figure 5.
The visual phenotype and radicle length of pea seeds treated with water (W), 0.3 M mannitol (M), and fullerol of different concentrations dissolved in 0.3 M mannitol (MF). MF1, MF2, MF3, and MF4 indicated fullerol dissolved in 0.3 M mannitol at the concentration of 10, 50, 100, and 500 mg/L, respectively. (a) One hundred and fifty grains of pea seeds each were used for phenotype analysis at 72 h after treatment. Radicle lengths were measured using a ruler in three replicates R1, R2, and R3 in all the treatments. (b) Multiple comparison results determined using the SSR-Test method were shown with lowercase letters to indicate statistical significance (P < 0.05).
Figure 6.
Venn diagram showing the shared and unique differentially expressed PsAQP genes in imbibing seeds under control (W), Mannitol (M) and Mannitol + Fullerol (MF3) treatments. Up-regulation (UG): PsPIP2-5, PsNIP1-1, PsNIP2-1-2, PsNIP4-2, PsNIP6-3, PsNIP1-5, PsTIP2-2, PsTIP4-1, PsSIP1-3, PsXIP2-1; Down-regulation (DG): PsTIP2-3, PsTIP3-2, PsNIP1-7, PsNIP5-1, PsXIP2-1.
Validation of the DEGs through qRT-PCR
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As a validation of the RNA-Seq data, eight genes showing differential expressions in imbibing seeds under M or M + F treatments were selected for qRT-PCR analysis, which was PsTIP4-1, PsTIP2-2, PsTIP2-3, PsTIP3-2, PsPIP2-5, PsXIP2-1, PsNIP6-3 and PsNIP1-5 shown in Fig 6, the expression modes of all the selected genes but PsXIP2-1 were well consistent between the RNA-Seq and the qRT-PCR data. PsXIP2-1, exhibiting slightly decreased expression under M treatment according to RNA-Seq, was found to be up-regulated under the same treatment by qRT-PCR (Fig. 7). This gene was therefore removed from further discussions.
Figure 7.
The expression patterns of seven PsAQPs in imbibing seeds as revealed by RNA-Seq and qRT-PCR. The seeds were sampled after 12 h soaking in three different solutions, namely water (W), 0.3 M mannitol (M), and 100 mg/L fullerol dissolved in 0.3 M mannitol (MF3) solution. Error bars are standard errors calculated from three replicates.
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A total of 39 full-length AQP genes belonging to five sub-families were identified from the pea genome and characterized for their sequences, phylogenetic relationships, gene structures, subcellular localization, and expression profiles. The number of AQP genes in pea is similar to that in related diploid legume species. The RNA-seq data revealed that PsTIP (2-3, 3-2) showed high expression in seeds for 5 d after imbibition, indicating their possible role during the initial phase of seed germination. Furthermore, gene expression profiles displayed that higher expression of PsTIP (2-3, 3-2) in germinating seeds might help maintain water balance under osmotic stress to confer tolerance. Our results suggests that the biological functions of fullerol in plant cells are exerted partly through the interaction with AQPs.
Deposition of raw data
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Under Bio project ID PRJNA793376 at the National Center for Biotechnology Information, raw data of sequencing read has been submitted. The accession numbers for the RNA-seq raw data are stored in GenBank and are mentioned in Supplemental Table S4.
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About this article
Cite this article
Pandey AK, Sun T, Wu X, Wang Z, Jiang R, et al. 2023. Aquaporin genes in garden pea and their regulation by the nano-antioxidant fullerol in imbibing embryos under osmotic stress. Vegetable Research 3:10 doi: 10.48130/VR-2023-0010
Aquaporin genes in garden pea and their regulation by the nano-antioxidant fullerol in imbibing embryos under osmotic stress
- Received: 22 November 2022
- Accepted: 16 January 2023
- Published online: 14 March 2023
Abstract: Aquaporins (AQPs) are known as small membrane intrinsic proteins that help to transport water and certain solutes through biological membranes. AQPs gene families have been extensively studied in major crops, but less investigated in pea (Pisum sativum L.), which is an economically significant legume crop with a huge complex genome. Here, we present a genome-wide identification, structural characterization, subcellular localization, and expression profiling of the AQPs in pea with a particular interest in their involvement in nano fullerol-conferred osmotic stress alleviation. We identified 39 full-length aquaporin genes from the pea genome, which were classified into five subfamilies. The protein structure of aquaporins appears to have substrate-specific residues which are conserved in plants, allowing for inference of substrate specificity. In particular, PsNIP2-2-2 was identified with a Gly-Ser-Gly-Arg (GSGR) selective filter that indicates the ability to uptake silicon. Analysis of tissue transcriptomes revealed preferred expressions of certain PsAQPs in the underground, aerial and reproductive organs, respectively. Development-regulated expression of two PsTIPs and two PsPIPs in seeds were noticed. RNA-Seq of the imbibing embryos treated with mannitol (M) or mannitol plus 100 mg/L fullerol (MF) revealed two PsTIPs being similarly regulated by M or MF, three PsNIPs being up-regulated only by M without F, and four other genes that were only regulated under MF condition. To our knowledge, this is the first report on transcriptional regulation of AQPs by fullerols, which adds to our knowledge on the plant-carbon nano-substances interactions.
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Key words:
- Membrane intrinsic proteins (MIPs) /
- Fullerol /
- Osmotic stress /
- Pea /
- Gene expression