Comparative proteomics analysis of two maize hybrids revealed drought-stress tolerance mechanisms

Figures & data

Figure 1. Phenotypic characterization of ears of the two maize hybrids. (a, b) Ear phenotypes. (c) Ear length. (d) Ear bare tip length. (e) Kernel rows per ear. (f) Kernels per row.

Figure 2. Number and grouping of DAPs. (a) Histogram of the number distribution of DAPs in four comparison groups. (b) Venn diagram analysis of DAPs identified in the four experimental comparisons. (c) Cluster analysis of DAPs in SD_TD comparison.

Table 1. DAPs observed specifically in tolerant hybrid ND476.

No.	Protein ID	Gene name/ID	Description	Fold change	p value	Regulated type	KEGG pathway
1	B6TF92	100283176	Stress responsive protein	1.30	2.11E-02	Up
2	A0A1D6MD20	Zm00001d039045	Ubiquitin carboxyl-terminal hydrolase 3	1.22	2.17E-02	Up
3	A0A1D6MBB0	542588	Nucleosome assembly protein 1	0.82	7.72E-03	Down
4	K7TR40	Zm00001d024640	Oxidoreductase family protein	1.21	3.28E-02	Up
5	A0A1D6FFF3	Zm00001d008779	Leucine-rich repeat extensin-like protein 3	0.78	3.31E-02	Down
6	A0A1D6KNW3	Zm00001d032166	Protein enhanced disease resistance	1.24	2.53E-02	Up
7	A0A1D6G3P7	100281761	Methylsterol monooxygenase 2-2	1.22	1.71E-02	Up	Steroid biosynthesis
8	B6T5W5	100283837	Nucleolin	1.22	8.42E-03	Up
9	A0A1D6F4V9	Zm00001d007215	26S proteasome regulatory subunit 4 homolog A	0.82	1.64E-03	Down	Proteasome
10	C4J608	100501797	Transcription initiation factor TFIID subunit 5	0.80	4.09E-02	Down	Basal transcription factors
11	A0A1D6KDZ6	Zm00001d030690	Translocon-associated protein beta (TRAPB) family protein	0.83	1.64E-02	Down	Protein processing in endoplasmic reticulum
12	B6SHX0	100286270	5a2 protein	1.57	3.40E-02	Up
13	A0A1D6MM68	Zm00001d040014	Nuclease PA3	1.30	1.39E-02	Up
14	A0A1D6FGJ9	Zm00001d008898	Peroxidase	1.31	3.64E-02	Up	Phenylpropanoid biosynthesis
15	J7LC26	Zm00001d042023	Ribosomal protein S10	0.83	4.46E-02	Down	Ribosome
16	B4F929	100272373	Carboxypeptidase	0.83	1.64E-02	Down
17	B6SGF4	100280457	Nonspecific lipid-transfer protein	1.64	2.39E-02	Up
18	Q94IR1	542066	Methyl binding domain101a	1.27	2.47E-02	Up
19	A0A1D6FJH2	Zm00001d009445	Double-stranded DNA-binding family protein	1.47	1.12E-02	Up
20	A0A1D6EM72	Zm00001d005377	Lecithin-cholesterol acyltransferase-like 1	0.74	4.39E-02	Down

Table 2. DAPs showed differentially expressed in both the tolerant and sensitive hybrids after drought treatment.

No.	Protein ID	Gene name/ID	Description	TD/TC Fold change	p value	TD/SD Fold change 4	p value	Regulated type	KEGG pathway
1	K7UBM6	Zm00001d053855	DNA binding protein isoform 1	1.21	1.18E-02	1.21	3.52E-03	Up
2	B6UI58	100279135	Uncharacterized protein	1.26	3.94E-02	1.23	1.78E-02	Up
3	B6SHM3	100274719	Uncharacterized protein	1.72	4.30E-02	1.92	4.36E-02	Up
4	B4FNB6	100277277	Mediator-associated protein 2	0.80	3.05E-02	0.77	2.17E-02	Down
5	C0PK55	100383938	Cytochrome c-2	1.22	1.32E-02	1.21	2.40E-02	Up
6	A0A1D6NL34	Zm00001d044342	40S ribosomal protein S23-2	0.67	2.06E-03	0.69	1.01E-02	Down
7	P27788	FDX3	Ferredoxin-3	1.31	7.32E-03	1.29	2.35E-02	Up	Photosynthesis
8	A0A1D6ESA9	100283408	DNA binding protein	1.28	2.22E-03	1.32	7.16E-03	Up
9	A0A1D6NTW2	Zm00001d045127	Uncharacterized protein	1.25	3.15E-02	1.35	3.91E-02	Up	Other types of O-glycan biosynthesis
10	A0A1D6IBP5	Zm00001d021507	Asparagine-tRNA ligase chloroplastic/mitochondrial	0.64	4.42E-02	0.67	4.98E-02	Down	Aminoacyl-tRNA biosynthesis
11	A0A1D6I963	100274090	Plant UBX domain-containing protein 8	2.96	2.91E-02	2.43	4.03E-02	Up
12	A0A1D6G380	Zm00001d011710	Cytidine deaminase 1	1.24	4.62E-02	1.41	2.85E-02	Up	Pyrimidine metabolism
13	A0A1D6INR0	Zm00001d022516	Stress responsive protein	1.45	2.56E-03	1.78	4.91E-02	Up

Table 3. Common (overlapping) drought-responsive DAPs in filling seed between ND476 and ZX 978.

No.	Protein ID	Gene name/ID	Description	TD/TC Fold Change	TD/TC Regulated type	TD/TC	SD/SC Fold change	SD/SC Regulated type	SD/SC	KEGG pathway
						p value			p value
1	C0PLU9	100281175	Hypoxanthine phosphoribosyltransferase	0.60	Down	1.58E-04	1.58	Up	1.17E-04	Purine metabolism
2	A0A1D6JWG9	Zm00001d028467	Uncharacterized protein	3.05	Up	8.10E-03	0.29	Down	1.36E-03
3	A0A1D6EAZ3	Zm00001d003698	CBS domain-containing protein CBSCBSPB2	2.32	Up	4.65E-04	0.38	Down	4.56E-04
4	A0A1D6FCB8	Zm00001d008380	rRNA processing protein-related	0.79	Down	4.10E-03	1.30	Up	4.95E-02

Table 4. Drought-responsive maize seedling leaf proteins observed specifically in sensitive hybrid ZX 978.

No.	Protein ID	Gene name/ID	Description	Fold change	p value	Regulated type	KEGG pathway
1	A0A1D6DT14	Zm00001d001808	Uncharacterized protein	1.42	3.71E-02	Up
2	A0A1D6L1L9	Zm00001d033683	Phospholipase A2	1.21	2.73E-02	Up	alpha-Linolenic acid metabolism
							Arachidonic acid metabolism
							Linoleic acid metabolism
							Ether lipid metabolism
							Glycerophospholipid metabolism
3	A0A1D6GLD8	Zm00001d013667	Methyl-CpG-binding domain-containing protein 11	1.26	3.25E-02	Up	Phenylpropanoid biosynthesis
4	A0A1D6K6A8	Zm00001d029611	Glycosyltransferase	1.31	3.30E-03	Up	Other types of O-glycan biosynthesis
5	B6SKJ7	100280687	Early nodulin-like protein 3	1.43	2.64E-02	Up
6	B6TAE3	101027138	TPR domain containing protein	1.27	3.10E-02	Up
7	K7TZ57	103641028	Uncharacterized protein	0.52	4.62E-03	Down
8	P30755	—	Histone H2B.1	1.20	3.34E-02	Up
9	A0A1D6HNZ3	100285265	Casein kinase I	0.81	2.46E-02	Down
10	B4F7T6	100191150	SelT-like protein	1.20	4.33E-02	Up
11	A0A1D6IA06	100284381	Protein binding protein	1.24	2.12E-02	Up
12	A0A1D6MWD1	Zm00001d041475	Septin and tuftelin-interacting protein 1 homolog 1	1.25	8.44E-03	Up
13	A0A1D6PXC5	103653151	Nucleic acid binding	1.44	4.92E-02	Up	Spliceosome
14	A0A1D6KW47	103643608	Protein DETOXIFICATION	0.83	1.08E-02	Down
15	A0A1D6JRN4	100281926	Sulfhydryl oxidase	0.83	1.72E-03	Down
16	B6SVH1	100275560	Uncharacterized protein	1.21	3.47E-02	Up
17	K7TYT2	Zm00001d048824	Putative oligouridylate binding protein	1.21	2.40E-02	Up
18	B6UGI6	100304071	Uncharacterized protein	1.24	2.46E-02	Up
19	B4FLF6	Zm00001d020765	DNA-damage-repair/toleration protein DRT111 chloroplastic	1.21	8.74E-03	Up	Spliceosome
20	A0A1D6KL36	BAG1_3	BAG family molecular chaperone regulator 1	1.22	2.09E-02	Up
21	A0A1D6GWE1	Zm00001d014792	Shepherd-like1	1.27	1.76E-03	Down	Plant-pathogen interaction
							Protein processing in endoplasmic reticulum

Figure 3. GO functional classification of drought responsive proteins. (a) ND 476 specific DAPs. (b) ZX 978 specific DAPs. The y-axis represents the number of proteins in each function; x-axis displays the protein functions, categorized into three broad biological functional groups.

Figure 4. KEGG pathway enrichment analysis of DAPs. (a) Most significantly enriched pathways in TD_TC. (b) Most significantly enriched pathways in SD_SC. The size of the point represents the number of DAPs. TC, tolerant cultivar (ND476) under control conditions; TD, tolerant cultivar under drought conditions; SC, sensitive cultivar (ZX978) under well-watered (control) conditions; SD, sensitive cultivar under drought treatment conditions. An underscore between two genotype-treatment combinations represents comparison between those combinations, for example, TD_TC.

Figure 5. Confirmation of the iTRAQ-seq results by qRT-PCR. (a) DAPs specific to TD_TC; (b) DAPs specific to SD_TD; (c) DAPs shared between TC_TD and SD_SC; and (d) Common DAPs shared between TD_TC and TD_SD. The y-axis represents expression levels (fold change). Maize gene GAPDH (accession no. X07156) was used as the internal reference. Error bars represent the SE (n = 3). TC, tolerant cultivar (ND 476) under control conditions; TD, tolerant cultivar under drought conditions; SC, sensitive cultivar (ZX 978) under well-watered (control) conditions; SD, sensitive cultivar under drought treatment conditions. An underscore between two genotype-treatment combinations represent comparison between those combinations, for example, TD_TC.