| Water is an ubiquitous and indispensable molecule for plant growth and development.According to the composite water flux model, water passes plant organs by three pathways:apoplastic, symplastic, and transcellular. The latter two routes can not be experimentallydistinguished and are combined as the cell-to-cell pathway. Aquaporins represent an importantselective pathway for water to move across cellular membranes. Genome studies have revealedthat aquaporins constitute a superfamily in plants. Arabidopsis. thaliana, Zea mays and Oryza sativahave 38, 35 and 33 genes, respectively. Based on amino-acid sequence homology andsubcellular location, plant aquaporins are classified into four subfamilies: PIPs for plasmamembrane intrinsic proteins;TIPs for tonoplast intrinsic proteins;NIPs for NOD26-likeintrinsic proteins;SIPs for small basic intrinsic proteins. The PIP subfamily has been furtherdivided into two major groups: PIP1 and PIP2. At the sequence level, PIP1s can bedistinguished from PIP2s proteins by a longer N-terminal extension and a shorter C-terminalend. PIP1s proteins exhibit no or very low water channel activity compared with PIP1sproteins, when expressed in the X. laevis oocyte heterologous system. Plant aquaporinactivity might be regulated by gating mechanisms. The factors affecting the gating behaviourpossibly involve phosphorylation, heteromerization, pH, Ca2+, osmotic pressure, solutegradients and temperature. Plant aquaporins are involved in seed germination, cellelongation, stomatal movement, fertilization and so on. Some plant aquaporins also play animportant role in response to stress. Many studies have focused on the relationship betweenplant aquaporins and stress resistance. Plants counteract fluctuations in water supply byregulating all aquaporins in the cell plasma membrane. An endemic drought-tolerant plant,Glycyrrhiza uralensis F., growing in the sandy desert at the west of China was collected andcultured for the molecular cloning of aquaporin genes and for monitoring the distribution andexpression patterns of aquaporins in order to further elucidate their involvement in eremicplant water transport.1. Two degenerate oligonucleotide primers were designed based on two conserved domainNPA boxes in the plant aquaporin amino acid sequence. Using RT-PCR, 5′RACE and3′RACE, a full length cDNA of aquaporin-like gene was isolated from cDNA preparedfrom Glycyrrhiza uralensis F. roots, named GuPIP1. The analysis of the sequence showsthat the GuPIP1 cDNA consists of 1,236-bp upstream of the poly (A) tail, which includes a71-bp 5′-noncoding region , followed by 870 bp of open reading frame encoding 290amino acids, and, finally, a 295-bp 3′-noncoding region (accession no. AY781788).2. The analysis of the amino acid sequence reveals that GuPIP1 contains the MIP familysequence SGXHXNPAVT, the PIP family signature sequence GGGANXXXXGY andTGI /TNPARSI/FGAAI/VI/VF/YN. GuPIP1 exhibits six predicted transmembrane helicesand possesses two sets of conserved NPA motifs as well as the highly conserved motifs onM1 and M4 as EXXXTXXF/L which may be involved in the recognition and transport ofwater molecules across the water channel. As compared to PIPs identified from the otherplant species AQPs, GuPIP1 has the highly sequence identity at the amino acid levelwith PIP1s among all plant AQPs compared, including AtPIP1;4 and ZmPIP1;5 testifiedas functional aquaporins. This aquaporin has a longer N-terminal extension thanC-terminal extension by 46 amino acids, which is consistent with the special PIP1signature sequence. So GuPIP1 should be a member of PIP1 subfamily.3. GuPIP1 protein was confirmed to be localized in the PM by immunochemical reaction withpeptide-specific antibodies. GuPIP1 was localized in epidermis cells and calyptrogen cells ofGlycyrrhiza uralensis F. radicles through immunohistochemical method. Western blotanalysis indicated that the expression of GuPIP1 was not tissue-specific but in roots, stems andleaves. So, GuPIP1 plays an important role in whole plant water transport.4. Quantitative real-time PCR analysis showed that the expression of GuPIP1 was up-regulated bydrought, ABA and salt stress. GuPIP1 may play a putative role as osmotic or turgor sensors.
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