Genome-wide analysis of metallothionein gene family in maize to reveal its role in development and stress resistance to heavy metal.

BACKGROUND: Maize (Zea mays L.) is a widely cultivated cereal and has been used as an optimum heavy metal phytoremediation crop. Metallothionein (MT) proteins are small, cysteine-rich, proteins that play important roles in plant growth and development, and the regulation of stress response to heavy metals. However, the MT genes for maize have not been fully analyzed so far. METHODS: The putative ZmMT genes were identified by HMMER.The heat map of ZmMT genes spatial expression analysis was generated by using R with the log2 (FPKM + 1).The expression profiles of ZmMT genes under three kinds of heavy metal stresses were quantified by using qRT-PCR. The metallothionein proteins was aligned using MAFFT and phylogenetic analysis were constructed by ClustalX 2.1. The protein theoretical molecular weight and pI, subcellular localization, TFs binding sites, were predicted using ProtParam, PSORT, PlantTFDB, respectively. RESULTS: A total of 9 ZmMT genes were identified in the whole genome of maize. The results showed that eight of the nine ZmMT proteins contained one highly conserved metallothio_2 domain, while ZmMT4 contained a Metallothio_PEC domain. All the ZmMT proteins could be classified into three major groups and located on five chromosomes. The ZmMT promoters contain a large number of hormone regulatory elements and hormone-related transcription factor binding sites. The ZmMT genes exhibited spatiotemporal specific expression patterns in 23 tissues of maize development stages and showed the different expression patterns in response to Cu, Cd, and Pb heavy metal stresses. CONCLUSIONS: We identified the 9 ZmMT genes, and explored their conserved motif, tissue expression patterns, evolutionary relationship. The expression profiles of ZmMT genes under three kinds of heavy metal stresses (Cu, Cd, Pb) were analyzed. In summary, the expression of ZmMTs have poteintial to be regulated by hormones. The specific expression of ZmMTs in different tissues of maize and the response to different heavy metal stresses are revealed that the role of MT in plant growth and development, and stress resistance to heavy metals.

Genome-wide analysis of metallothionein gene family in maize to reveal its role in development and stress resistance to heavy metal

BACKGROUND: Maize (Zea mays L.) is a widely cultivated cereal and has been used as an optimum heavy metal phytoremediation crop. Metallothionein (MT) proteins are small, cysteine-rich, proteins that play important roles in plant growth and development, and the regulation of stress response to heavy metals. However, the MT genes for maize have not been fully analyzed so far. METHODS: The putative ZmMT genes were identified by HMMER. The heat map of ZmMT genes spatial expression analysis was generated by using R with the log2 (FPKM + 1). The expression profiles of ZmMT genes under three kinds of heavy metal stresses were quantified by using qRT-PCR. The metallothionein proteins was aligned using MAFFT and phylogenetic analysis were constructed by ClustalX 2.1. The protein theoretical molecular weight and pI, subcellular localization, TFs binding sites, were predicted using ProtParam, PSORT, PlantTFDB, respectively. RESULTS: A total of 9 ZmMT genes were identified in the whole genome of maize. The results showed that eight of the nine ZmMT proteins contained one highly conserved metallothio_2 domain, while ZmMT4 contained a Metallothio_PEC domain. All the ZmMT proteins could be classified into three major groups and located on five chromosomes. The ZmMT promoters contain a large number of hormone regulatory elements and hormone-related transcription factor binding sites. The ZmMT genes exhibited spatiotemporal specific expression patterns in 23 tissues of maize development stages and showed the different expression patterns in response to Cu, Cd, and Pb heavy metal stresses. CONCLUSIONS: We identified the 9 ZmMT genes, and explored their conserved motif, tissue expression patterns, evolutionary relationship. The expression profiles of ZmMT genes under three kinds of heavy metal stresses (Cu, Cd, Pb) were analyzed. In summary, the expression of ZmMTs have poteintial to be regulated by hormones. The specific expression of ZmMTs in different tissues of maize and the response to different heavy metal stresses are revealed that the role of MT in plant growth and development, and stress resistance to heavy metals.

Spermidine Suppressed the Inhibitory Effects of Polyamines Inhibitors Combination in Maize (Zea mays L.) Seedlings under Chilling Stress.

Chilling stress greatly inhibited the seed germination, plant growth, development and productivity in this study. The current research aimed to study the effects of different polyamine (PA) inhibitor combinations (Co), e.g., D-arginine (D-Arg), difluoromethylormithine (DFMO), aminoguanidine (Ag) and methylglyoxyl-bis-(guanyhydrazone) (MGBG) at different doses, i.e., 10 µM Co, 100 µM Co, 500 µM Co, 1000 µM Co and 1000 µM Co + 1 mM Spd (Spermidine) in two inbred lines of maize (Zea mays L.), i.e., Mo17 and Huang C, a sensitive and tolerant chilling stress, respectively. The combination treatments of PA inhibitors reduced the biosynthesis of putrescine (Put) in the tissues of both studied inbred lines. Application with 500 µM Co and 1000 µM Co did not result in a significant difference in Put concentrations, except in the coleoptile of Mo17. However, combining Spd to 1000 µM of PA inhibitors enhanced the Put, Spd, spermine (Spm) and total PAs in the roots, coleoptile and mesocotyls. Put and total PAs were increased by 39.7% and 30.54%, respectively, when Spd + 1000 µM Co were applied relative to their controls. Chilling stress and PA inhibitors treatments affected both inbred lines and resulted in differences in the PA contents. Results showed that enzymes involved in the biosynthesis of PAs (ornithine decarboxylase as ODC and S-adenosylmethionine decarboxylase as SAMDC) were significantly downregulated by 1000 µM Co in the tissues of both inbred lines. In contrast, the activity of PAO, a Pas degradation enzyme, was significantly improved by 1000 µM Co under chilling stress. However, Spd + 1000 µM Co significantly improved the activities of ODC and SAMDC and their transcript levels (ODC and SAMDC2). While it significantly downregulated the PAO activity and their relative genes (PAO1, PAO2 and PAO3) under chilling stress. Overall, this study elucidates the specific roles of Spd on the pathway of PA inhibitors and PA biosynthesis metabolism in maize seed development in response to chilling stress. Moreover, the Huang C inbred line was more tolerant than Mo17, which was reflected by higher activities of PA biosynthesis-related enzymes and lower activities of PAs' degradative-related enzymes in Huang C.

Polyamine biosynthetic pathways and their relation with the cold tolerance of maize (Zea mays L.) seedlings.

BACKGROUND: The present study was designed to investigate the inhibition role of two polyamine biosynthesis inhibitors, i.e., D-arginine (D-Arg) and DL-α-difluoromethylornithine (DFMO), in polyamine biosynthesis under chilling stress in different tissues of two maize inbred lines - Huang C (chilling-tolerance) and Mo17 (chilling-sensitive). RESULTS: The results showed that exposure to the lower concentration of polyamine biosynthesis inhibitors improved seedlings growth, such as the root length, root and shoot fresh weight, chlorophyll a (chl a). The effectiveness of 10 µM D-Arg treatments was more prominent than those of 10 µM DFMO. However, the higher concentration of inhibitors suppressed seedlings growth, and the exposure to 100 µM DFMO caused stronger decreases in the photosynthetic pigments, such as chlorophyll a (chl a), chlorophyll b (chl b), total chlorophyll and carotenoids, than the other treatments. Meanwhile, the inhibitor treatments caused the lower content of putrescine (Put) in roots, mesocotyls and coleoptiles in both maize inbred lines as compared with untreated plants. However, the lower concentration (10 µM) of polyamine biosynthetic inhibitors improved the Spd content, except 10 µM D-Arg in root of Huang C, and 10 µM DFMO in coleoptiles of both Mo17 and Huang C. The correlation analysis found that Spd was positively significantly correlated with root length and shoot fresh weight of seedling. CONCLUSION: It was showed that the Spd played an important role in seedling growth improvement. At the same concentration of polyamine biosynthetic inhibitors, the Put contents in different tissues of the seedlings treated with DFMO were generally lower than those treated with D-Arg, except for Put contents in root of Mo17 with 10 µM treatment. Moreover, the treatments of 100 µM were more prominent than those of 10 µM treatments. Exposure to 100 µM D-Arg and 100 µM DFMO could each decrease the activities of Arginine decarboxylase (ADC), Ornithine decarboxylase (ODC) and S-adenosylmethionine decarboxylase (SAMDC) in all maize tissues. However, the decrease of the ADC activity was more prominent in 100 µM D-Arg-treated seedlings, while the decrease of SAMDC and ODC activities was prominent in 100 µM DFMO-treated seedlings. Genes involved in polyamine biosynthesis, such as ADC, ODC, SAMDC, and PAO, showed different expression patterns in response to chilling stress and polyamine biosynthesis inhibitors. This study suggested that Put was synthesized via both the ADC and ODC pathways after chilling stress, with the ODC pathway being the major one.

Maize annexin genes ZmANN33 and ZmANN35 encode proteins that function in cell membrane recovery during seed germination.

Plasma membrane (PM) recovery from the impaired dry state is essential for seed germination, but its underlying mechanism remains unclear. In this study, we found that ZmANN33 and ZmANN35, two annexin genes in maize, encode proteins that participate in PM recovery during seed germination. The expression of both genes was up-regulated during seed germination and strongly repressed by chilling (either 15 or 5 °C) as compared with the normal temperature (25 °C). In addition, the increased membrane damage caused by chilling imbibition was correlated with suppressed expression of ZmANN33 and ZmANN35, while rapid recovery of their expression levels accompanied the rescue of the damaged membrane. Arabidopsis seedlings ectopically expressing ZmANN33 or ZmANN35 had longer seedling length than wild-type (WT) plants during the recovery period after 3 d of chilling stress, indicating the positive roles of these two gene products in the plant's recovery from chilling injury. Moreover, these transgenic seedlings had lower lipid peroxidation and higher peroxidase activities than WT during the recovery period. Consistently, root cells of these transgenic seedlings had more intact PM after chilling stress, supporting the proposition that ZmANN33 and ZmANN35 contribute to the maintenance of PM integrity. The enhanced PM integrity is likely due to the accelerated exocytotic process after chilling stress. We also showed that both ZmANN33 and ZmANN35 localized in the cytosol near the plasma membrane. Thus, we conclude that ZmANN33 and ZmANN35 play essential roles in membrane recovery during maize seed germination.

[Antioxidant enzyme activities and proline content in maize seedling and their relationships to cold endurance].

This paper studied the catalase (CAT) and peroxidase (POD) activities and proline (Pro) content in the seedling's root, mesocotyl (including coleoptile node) , and coleoptile (including infancy leaf) of two maize inbred lines Huang C (cold-endurance) and Mo17 (cold-sensitive), and their relationships to cold endurance under low temperature stress (5 degrees C, 3 d). The results showed that under the low temperature stress, Huang C had a significantly higher relative growth rate of mesocotyl and coleoptile than Mo17, and the change rates of CAT and POD activities and Pro content in the root, mesocotyl and coleoptile of Huang C were higher than those of Mo17. The plasma membrane permeability of the three segments of both inbred lines was significantly increased, but Huang C had significantly lower plasma membrane permeability than Mo17. Compared with those under normal temperature, the seedlings under low temperature stress had higher CAT activity and Pro content, and the increment was higher in Huang C than in Mo17. The POD activity in the three segments of the seedlings under low temperature stress was reduced, and the decrement was lower in Huang C than in Mo17. Stepwise regression analysis showed that under low temperature stress, the dry matter accumulation in cold-endurance line Huang C was mainly related to Pro content, while that in cold-sensitive line Mo17 was mainly related to POD activity. Mesocotyl was the most responsive tissue to low temperature. The change rate of Pro content in mesocotyl was more closely correlated with the cold endurance of maize inbred lines than the other physiological parameters.