Identification and Functional Characterization of Ammonia transporter Genes <i>PdRhp-1</i> in <i>Pocillopora damicornis</i>

Zeng Sainan; Qin Zhixuan; Wang Yi; Liu Zhaoqun; Zhou Zhi

doi:10.12284/hyxb2025124

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Zeng Sainan，Qin Zhixuan，Wang Yi, et al. Identification and Functional Characterization of Ammonia transporter Genes PdRhp-1 in Pocillopora damicornis[J]. Haiyang Xuebao，2025, 47(11)：1–11 doi: 10.12284/hyxb2025124

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Zeng Sainan，Qin Zhixuan，Wang Yi, et al. Identification and Functional Characterization of Ammonia transporter Genes PdRhp-1 in Pocillopora damicornis[J]. Haiyang Xuebao，2025, 47(11)：1–11 doi: 10.12284/hyxb2025124

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Identification and Functional Characterization of Ammonia transporter Genes PdRhp-1 in Pocillopora damicornis

doi: 10.12284/hyxb2025124

Zeng Sainan^1
,,
Qin Zhixuan²,
Wang Yi¹,
Liu Zhaoqun^{2
,
,},
Zhou Zhi^{2
,
,}

1.
Hainan University, School of Marine Biology and Fisheries, Haikou 570228, China
2.
Hainan University, School of Marine Science and Engineering, Haikou 570228, China

Received Date: 2025-06-08
Rev Recd Date: 2025-07-24

Available Online: 2025-08-05

Abstract

Abstract

To reveal the effect of high temperature on the ammonia assimilation of Pocillopora damicornis and elucidate the thermal adaptation mechanism of scleractinian coral, we identified and cloned an ammonia transporter gene PdRhp-1 from P. damicornis. The open reading frame (ORF) of PdRhp-1 is 1410 bp, encoding a polypeptide chain composed of 469 amino acid residues. Sequence analysis showed that PdRhp-1 was a hydrophobic transmembrane protein, containing 12 transmembrane domains, belonging to the Rhesus-type ammonia transporter. Its amino acid sequence shares 44.14% identity with Homo sapiens ammonia transporter gene RhCG. In order to analyze the biological function of PdRhp-1, its recombinant expression vector was transfected into HEK293T cells, and ammonium chloride was added to the culture medium. It was found that the total ammonia uptake rate in the cells expressing PdRhp-1 was significantly higher than that in the control group, indicating that PdRhp-1 has ammonia transport function. At the same time, the transcriptome data of high temperature treated P. damicornis were analyzed, and it was found that high temperature significantly inhibited the expression of PdRhp-1 and some genes related to ammonia assimilation. The above results demonstrate that PdRhp-1 is a Rhesus-type ammonia transporter, and high temperature may affect the symbiotic homeostasis of corals and zooxanthellae by inhibiting the ammonia transport process mediated by PdRhp-1.
- Pocillopora damicornis,
- ammonia transporter,
- ammonia assimilation,
- high temperature

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References

[1]	Fisher R, O’Leary R A, Low-Choy S, et al. Species richness on coral reefs and the pursuit of convergent global estimates[J]. Current Biology, 2015, 25(4): 500−505. doi: 10.1016/j.cub.2014.12.022
[2]	Graham N A J, Nash K L. The importance of structural complexity in coral reef ecosystems[J]. Coral Reefs, 2013, 32(2): 315−326. doi: 10.1007/s00338-012-0984-y
[3]	Ortiz D M, Tissot B N. Evaluating ontogenetic patterns of habitat use by reef fish in relation to the effectiveness of marine protected areas in West Hawaii[J]. Journal of Experimental Marine Biology and Ecology, 2012, 432-433: 83-93.
[4]	McLachlan R H, Dobson K L, Schmeltzer E R, et al. A review of coral bleaching specimen collection, preservation, and laboratory processing methods[J]. PeerJ, 2021, 9: e11763. doi: 10.7717/peerj.11763
[5]	Cleves P A, Krediet C J, Lehnert E M, et al. Insights into coral bleaching under heat stress from analysis of gene expression in a sea anemone model system[J]. Proceedings of the National Academy of Sciences, 2020, 117(46): 28906−28917. doi: 10.1073/pnas.2015737117
[6]	Qin Binni, Yu Kefu, Zuo Xiuling. Study of the bleaching alert capability of the CRW and CoRTAD coral bleaching heat stress products in China's coral reefs[J]. Marine Environmental Research, 2023, 186: 105939. doi: 10.1016/j.marenvres.2023.105939
[7]	Zuo Xiuling, Qin Binni, Teng Juncan, et al. Optimized spatial and temporal pattern for coral bleaching heat stress alerts for China's coral reefs[J]. Marine Environmental Research, 2023, 191: 106152. doi: 10.1016/j.marenvres.2023.106152
[8]	Pernice M, Meibom A, Van Den Heuvel A, et al. A single-cell view of ammonium assimilation in coral–dinoflagellate symbiosis[J]. The ISME Journal, 2012, 6(7): 1314−1324. doi: 10.1038/ismej.2011.196
[9]	李淑, 余克服. 珊瑚礁白化研究进展[J]. 生态学报, 2007, 27(5): 2059−2069. doi: 10.3321/j.issn:1000-0933.2007.05.047 Li Shu, Yu Kefu. Recent development in coral reef bleaching research[J]. Acta Ecologica Sinica, 2007, 27(5): 2059−2069. doi: 10.3321/j.issn:1000-0933.2007.05.047
[10]	Muscatine L, Porter J W. Reef corals: mutualistic symbioses adapted to nutrient-poor environments[J]. Bioscience, 1977, 27(7): 454−460. doi: 10.2307/1297526
[11]	Xiang Tingting, Lehnert E, Jinkerson R E, et al. Symbiont population control by host-symbiont metabolic interaction in Symbiodiniaceae-cnidarian associations[J]. Nature Communications, 2020, 11(1): 108. doi: 10.1038/s41467-019-13963-z
[12]	Lin Senjie, Cheng Shifeng, Song Bo, et al. The Symbiodinium kawagutii genome illuminates dinoflagellate gene expression and coral symbiosis[J]. Science, 2015, 350(6261): 691−694. doi: 10.1126/science.aad0408
[13]	Rädecker N, Pogoreutz C, Gegner H M, et al. Heat stress destabilizes symbiotic nutrient cycling in corals[J]. Proceedings of the National Academy of Sciences of the United States of America, 2021, 118(5): e2022653118.
[14]	Kemp D W, Hoadley K D, Lewis A M, et al. Thermotolerant coral–algal mutualisms maintain high rates of nutrient transfer while exposed to heat stress[J]. Proceedings of the Royal Society B: Biological Sciences, 2023, 290(2007): 20231403. doi: 10.1098/rspb.2023.1403
[15]	Tanaka Y, Suzuki A, Sakai K. The stoichiometry of coral-dinoflagellate symbiosis: carbon and nitrogen cycles are balanced in the recycling and double translocation system[J]. The ISME Journal, 2018, 12(3): 860−868. doi: 10.1038/s41396-017-0019-3
[16]	Wright P A. Nitrogen excretion: three end products, many physiological roles[J]. Journal of Experimental Biology, 1995, 198(2): 273−281. doi: 10.1242/jeb.198.2.273
[17]	Szmant A M, Ferrer L M, FitzGerald L M. Nitrogen excretion and O: N ratios in reef corals: evidence for conservation of nitrogen[J]. Marine Biology, 1990, 104(1): 119−127. doi: 10.1007/BF01313165
[18]	D'elia C F, Domotor S L, Webb K L. Nutrient uptake kinetics of freshly isolated zooxanthellae[J]. Marine Biology, 1983, 75(2): 157−167.
[19]	Miller D J, Yellowlees D. Inorganic nitrogen uptake by symbiotic marine cnidarians: a critical review[J]. Proceedings of the Royal Society of London. B. Biological Sciences, 1989, 237(1286): 109−125. doi: 10.1098/rspb.1989.0040
[20]	Khademi S, O'Connell III J, Remis J, et al. Mechanism of ammonia transport by Amt/MEP/Rh: structure of AmtB at 1.35 Å[J]. Science, 2004, 305(5690): 1587−1594. doi: 10.1126/science.1101952
[21]	Ludewig U, Von Wirén N, Frommer W B. Uniport of ${{\rm {NH}}_4^+} $ by the root hair plasma membrane ammonium transporter LeAMT1;1[J]. Journal of Biological Chemistry, 2002, 277(16): 13548−13555. doi: 10.1074/jbc.M200739200
[22]	Weiner I D, Verlander J W. Ammonia transport in the kidney by Rhesus glycoproteins[J]. American Journal of Physiology-Renal Physiology, 2014, 306(10): F1107−F1120. doi: 10.1152/ajprenal.00013.2014
[23]	Lupo D, Li Xiaodan, Durand A, et al. The 1.3-Å resolution structure of Nitrosomonas europaea Rh50 and mechanistic implications for NH₃ transport by Rhesus family proteins[J]. Proceedings of the National Academy of Sciences of the United States of America, 2007, 104(49): 19303−19308.
[24]	Huang Chenghan, Peng Jianbin. Evolutionary conservation and diversification of Rh family genes and proteins[J]. Proceedings of the National Academy of Sciences of the United States of America, 2005, 102(43): 15512−15517.
[25]	Westhoff C M, Ferreri-Jacobia M, Mak D O D, et al. Identification of the erythrocyte Rh blood group glycoprotein as a mammalian ammonium transporter[J]. Journal of Biological Chemistry, 2002, 277(15): 12499−12502. doi: 10.1074/jbc.C200060200
[26]	Chng Y R, Ong J L Y, Ching B, et al. Molecular characterization of three Rhesus glycoproteins from the gills of the African lungfish, Protopterus annectens, and effects of aestivation on their mRNA expression levels and protein abundance[J]. PLoS One, 2017, 12(10): e0185814. doi: 10.1371/journal.pone.0185814
[27]	张云龙, 张海龙, 王凌宇, 等. 氨氮对鱼类毒性的影响因子及气呼吸型鱼类耐氨策略[J]. 水生生物学报, 2017, 41(5): 1157−1168. doi: 10.7541/2017.144 Zhang Yunlong, Zhang Hailong, Wang Lingyu, et al. Impact factors of ammonia toxicity and strategies for ammonia tolerance in air-breathing fish: a review[J]. Acta Hydrobiologica Sinica, 2017, 41(5): 1157−1168. doi: 10.7541/2017.144
[28]	Huang Chenghan, Ye Mao. The Rh protein family: gene evolution, membrane biology, and disease association[J]. Cellular and Molecular Life Sciences, 2010, 67(8): 1203−1218. doi: 10.1007/s00018-009-0217-x
[29]	Lehnert E M, Mouchka M E, Burriesci M S, et al. Extensive differences in gene expression between symbiotic and aposymbiotic cnidarians[J]. G3 Genes\| Genomes\| Genetics, 2014, 4(2): 277−295.
[30]	Ganot P, Moya A, Magnone V, et al. Adaptations to endosymbiosis in a cnidarian-dinoflagellate association: differential gene expression and specific gene duplications[J]. PLoS Genetics, 2011, 7(7): e1002187. doi: 10.1371/journal.pgen.1002187
[31]	Capasso L, Zoccola D, Ganot P, et al. SpiAMT1d: molecular characterization, localization, and potential role in coral calcification of an ammonium transporter in Stylophora pistillata[J]. Coral Reefs, 2022, 41(4): 1187−1198. doi: 10.1007/s00338-022-02256-5
[32]	Thies A B, Quijada-Rodriguez A R, Zhouyao H, et al. A Rhesus channel in the coral symbiosome membrane suggests a novel mechanism to regulate NH₃ and CO₂ delivery to algal symbionts[J]. Science Advances, 2022, 8(10): eabm0303. doi: 10.1126/sciadv.abm0303
[33]	Yang Qingsong, Ling Juan, Zhang Ying, et al. Microbial nitrogen removal in reef-building corals: a light-sensitive process[J]. Chemosphere, 2024, 359: 142394. doi: 10.1016/j.chemosphere.2024.142394
[34]	Wang Ke, Chen Huiwen, Cheng Lili, et al. Structure and transport mechanism of human riboflavin transporters[J]. Nature Communications, 2025, 16(1): 4078. doi: 10.1038/s41467-025-59255-7
[35]	Liu Zhi, Peng Jianbin, Mo Rong, et al. Rh type B glycoprotein is a new member of the Rh superfamily and a putative ammonia transporter in mammals[J]. Journal of Biological Chemistry, 2001, 276(2): 1424−1433. doi: 10.1074/jbc.M007528200
[36]	Cunning R, Bay R A, Gillette P, et al. Comparative analysis of the Pocillopora damicornis genome highlights role of immune system in coral evolution[J]. Scientific Reports, 2018, 8(1): 16134. doi: 10.1038/s41598-018-34459-8
[37]	黄林韬, 黄晖, 江雷. 中国造礁石珊瑚分类厘定[J]. 生物多样性, 2020, 28(4): 515−523. doi: 10.17520/biods.2019384 Huang Lintao, Huang Hui, Jiang Lei. A revised taxonomy for Chinese hermatypic corals[J]. Biodiversity Science, 2020, 28(4): 515−523. doi: 10.17520/biods.2019384
[38]	高嵩钰. 海月水母(Aurelia coerulea)中主要毒素分子影响其变态发育的机制研究[D]. 上海: 中国人民解放军海军军医大学, 2022. Gao Songyu. Study on the mechanism of main toxin moleculesin Aurita coerulea affecting its metamorphosis[D]. Shanghai: Naval Medical University, 2022.
[39]	Hoadley K D, Lewis A M, Wham D C, et al. Host–symbiont combinations dictate the photo-physiological response of reef-building corals to thermal stress[J]. Scientific Reports, 2019, 9(1): 9985. doi: 10.1038/s41598-019-46412-4
[40]	Hoegh-Guldberg O. Climate change, coral bleaching and the future of the world's coral reefs[J]. Marine and Freshwater Research, 1999, 50(8): 839−866.
[41]	Yu Xiaopeng, Yu Kefu, Chen Biao, et al. Different responses of scleractinian coral Acropora pruinosa from Weizhou Island during extreme high temperature events[J]. Coral Reefs, 2021, 40(6): 1697−1711. doi: 10.1007/s00338-021-02182-y
[42]	Pfab F, Detmer A R, Moeller H V, et al. Heat stress and bleaching in corals: a bioenergetic model[J]. Coral Reefs, 2024, 43(6): 1627−1645. doi: 10.1007/s00338-024-02561-1
[43]	Lesser M P, Falcón L I, Rodríguez-Román A, et al. Nitrogen fixation by symbiotic cyanobacteria provides a source of nitrogen for the scleractinian coral Montastraea cavernosa[J]. Marine Ecology Progress Series, 2007, 346: 143−152. doi: 10.3354/meps07008
[44]	Gruswitz F, Chaudhary S, Ho J D, et al. Function of human Rh based on structure of RhCG at 2.1 Å[J]. Proceedings of the National Academy of Sciences of the United States of America, 2010, 107(21): 9638−9643.
[45]	Weiner I D, Verlander J W. Ammonia transporters and their role in acid-base balance[J]. Physiological Reviews, 2017, 97(2): 465−494. doi: 10.1152/physrev.00011.2016
[46]	Benghezal M, Gotthardt D, Cornillon S, et al. Localization of the Rh50-like protein to the contractile vacuole in Dictyostelium[J]. Immunogenetics, 2001, 52(3): 284−288.
[47]	Baday S, Orabi E A, Wang Shihao, et al. Mechanism of NH₄⁺ recruitment and NH₃ transport in Rh proteins[J]. Structure, 2015, 23(8): 1550−1557. doi: 10.1016/j.str.2015.06.010
[48]	Li X D, Lupo D, Zheng L, et al. Structural and functional insights into the AmtB/Mep/Rh protein family[J]. Transfusion Clinique et Biologique, 2006, 13(1/2): 65−69.
[49]	Marini A M, Boeckstaens M, Benjelloun F, et al. Structural involvement in substrate recognition of an essential aspartate residue conserved in Mep/Amt and Rh-type ammonium transporters[J]. Current Genetics, 2006, 49(6): 364−374. doi: 10.1007/s00294-006-0062-5
[50]	Benjelloun F, Bakouh N, Fritsch J, et al. Expression of the human erythroid Rh glycoprotein (RhAG) enhances both NH₃ and NH₄⁺ transport in HeLa cells[J]. Pflügers Archiv, 2005, 450(3): 155−167.
[51]	Bakouh N, Benjelloun F, Hulin P, et al. NH₃ is involved in the NH₄⁺ transport induced by the functional expression of the human Rh C glycoprotein[J]. Journal of Biological Chemistry, 2004, 279(16): 15975−15983. doi: 10.1074/jbc.M308528200
[52]	Dudler N, Miller D J. Characterization of two glutamate dehydrogenases from the symbiotic microalga Symbiodinium microadriaticum isolated from the coral Acropora formosa[J]. Marine Biology, 1988, 97(3): 427−430. doi: 10.1007/BF00397773