Edited by: Francisco Javier Romera, University of Córdoba, Spain
Reviewed by: Andriele Wairich, University of Bonn, Germany; Xin-Yuan Huang, Nanjing Agricultural University, China
This article was submitted to Plant Nutrition, a section of the journal Frontiers in Plant Science
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Network analysis is a systems biology-oriented approach based on graph theory that has been recently adopted in various fields of life sciences. Starting from mitochondrial proteomes purified from roots of
Living organisms are increasingly viewed as integrated and communicating molecular networks, thanks also to the diffusion of data-derived Systems Biology approaches (
Plant systems biology studies are dominated by transcriptomic data and statistics that, by measuring the dependence between variables (transcripts), allow us to reconstruct and analyze co-expression network models (
The co-expression networks reconstructed from proteomic data may represent an alternative to the lack of accurate PPI models and a tool for handling, at the system level, large-scale proteomic datasets related to the non-model plant. Similar to transcript networks, protein co-expression networks are defined as an undirected graph where nodes correspond to proteins and edges indicate significant correlation scores. By exploiting the laws underlying graph theory, the identification of hubs/bottlenecks and of differentially correlated proteins are sought, as well as the topological and functional modules related to specific biological phenotypes (
The studies that elucidate plant iron (Fe) nutrition are steadily increasing due to their impact on alleviating nutritional deficiencies in humans (
Given these premises, we performed the topological analysis of PPI and protein co-expression networks to identify new proteins involved in Fe and molybdenum (Mo) homeostasis in plants by taking into account subcellular compartmentalization. In particular, we reconstructed and analyzed networks from mitochondrial proteomes purified from roots of
A set of proteins differentially expressed in
Protein co-expression networks were reconstructed by processing
Both PPI and co-expression networks were topologically analyzed by Centiscape Cytoscape’s App (
The analysis of root mitochondria of
Fe starvation, combined with Mo starvation or sufficiency, upregulates several proteins involved in amino acid metabolism, such as glutamate dehydrogenases GDH1 and GDH2, aminobutyric acid transaminase POP2 involved in the metabolism of γ-amino butyrate (GABA), which links C and N metabolism in mitochondria (
The redox homeostasis group includes formate dehydrogenase (FDH), the enzyme catalyzing the oxidation of formate into CO2 (
The fatty acid metabolism group includes dihydrolipoamide branched chain acyltransferase BCE2, also known as Dark Inducible 3 (DIN3), whose expression increases soon after exposure to darkness (
The protein folding group includes the brassinosteroid (BR) biosynthesis gene DWF1, which mediates BR biosynthesis during the positive growth responses of the root system to low nitrogen (
The cell cycle/division group includes the mitochondrial GTPase MIRO1 (
The histone/chromatin group includes GYRB2, a DNA topoisomerase (
The translation group includes the pentatricopeptide protein PPR336, which associates with mitochondrial ribosomes (
The electron transport chain group includes the mitochondrial carrier UCP1/PUMP1 transporting aspartate
Transmembrane transporter activity group includes OM47 related to the voltage-dependent anion channel VDAC family; members of this family are major components of the outer mitochondrial membrane and are involved in the channeling of the products of chloroplast breakdown into the mitochondrion and in the exchange of various compounds between the cytosol and the mitochondrial intermembrane space (
The proteins downregulated under Fe or Mo starvation can represent interesting homeostatic regulators in the PPI networks, and their occurrence should not be neglected in future, more extensive analyses; as an example, rice OsIRO3 plays an important role in the Fe deficiency response by negatively regulating the
Co-expression networks were reconstructed from proteomes of purified mitochondria of
Proteins differentially correlated in
Annotation |
LDA |
Degree |
Function | |||||||||
UNIPROT ID ( |
Gene name |
Gene name |
Homology | Score | F Ratio | Prob > F | +Fe | +Mo | −Fe | −Mo | ||
A0A0A0K4Q8 | Csa_7G073600 | MCCB (AT4G34030) | 77% | 918 | 11,1 | 1,1E-04 | 5 | 2 | 9 | Methylcrotonyl-CoA carboxylase, subunit beta | ||
A0A0A0LM23 | Csa_2G382440 | FLOT1 (AT5G25250) | 75% | 723 | 4,6 | 1,2E-02 | Flotilins-like | |||||
A0A0A0LXD4 | Csa_1G424875 | AT4G30010 | 69% | 142 | 1 | 2 | 8 | ATP-dependent RNA helicase | ||||
A0A0A0L0B9 | Csa_4G082380 | MBL1 (AT1G78850) | 50% | 434 | 3,9 | 2,1E-02 | Mannose-binding lectin | |||||
A0A0A0K5L0 | Csa_7G387180 | MCCA (AT1G03090) | 69% | 1023 | 4 | 3 | 10 | Methyl crotonyl-CoA carboxylase subunit alpha | ||||
A0A0A0KJ29 | Csa_6G526470 | AT3G58140 | 74% | 657 | Phenylalanyl-tRNA synthetase | |||||||
A0A0A0KCX8 | Csa_6G077980 | ARGAH1 (AT4G08900) | 85% | 590 | 1 | 0 | 4 | Arginase | ||||
A0A0A0LBW6 | Csa_3G199630 | EDA9 (AT4G34200) | 83% | 981 | 4 | 1,9E-02 | 7 | D-3-phosphoglycerate dehydrogenase | ||||
A0A0A0L0I0 | Csa_4G285780 | PA2 (AT5G06720) | 53% | 335 | 3,7 | 2,6E-02 | 3 | 1 | 3 | Peroxidase | ||
A0A0A0KA81 | Csa_6G088110 | AT2G20420 | 88% | 757 | 4 | 8 | 12 | Succinyl-CoA ligase | ||||
A0A0A0LBB3 | Csa_3G760530 | SVL1 (AT5G55480) | 56% | 851 | 6,6 | 2,3E-03 | Glycerophosphoryl diester- phosphodiesterase | |||||
A0A0A0LFK3 | Csa_3G734240 | TIM9 (AT3G46560) | 82% | 162 | 0 | 2 | 1 | Translocase of the inner membrane 9 | ||||
A0A0A0KRD9 | Csa_5G613510 | UOX (AT2G26230) | 68% | 434 | Urate oxidase | |||||||
A0A0A0L542_ A0A0A0LXV7 | Csa_3G078260 Csa_1G660150 | GPT2 (AT1G61800) | 75% | 568 | 5 | 8,4E-03 | Glucose-6-phosphate/phosphate translocator | |||||
A0A0A0K1S9_ A0A0A0K3R7 | Csa_7G047450 Csa_7G047440 | AT2G20710 | 45% | 415 | 8 | 3 | 1 | PPR-type organelle RNA editing factor | ||||
A0A0A0LS81 | Csa_1G096620 | ASP1 (AT2G30970) | 88% | 768 | 1 | 11 | 0 | Aspartate transaminase | ||||
A0A0A0LQ27 | Csa_1G025890 | OAT (AT5G46180) | 79% | 738 | 7 | 3 | Ornithine delta aminotransferase | |||||
A0A0A0L404 | Csa_4G646110 | FTSH4 (AT2G26140) | 81% | 1128 | 2 | 1 | 1 | ATP-dependent zinc metalloprotease | ||||
A0A0A0KIM0 | Csa_6G497010 | NFS1 (AT5G65720) | 78% | 756 | 2 | Cysteine desulfurase | ||||||
A0A0A0KB82 | Csa_7G407690 | AT5G61310 | 67% | 95,1 | 4 | Cytochrome c oxidase subunit | ||||||
A0A0A0L3T5 | Csa_4G642530 | VDAC2 (AT5G67500) | 53% | 317 | Voltage-gated anion channel | |||||||
A0A0A0KMP0 | Csa_5G321480 | AT2G07698 | 93% | 562 | 12 | 15 | 1 | ATP synthase subunit alpha | ||||
A0A0A0LGF5 | Csa_2G033990 | LON1 (AT5G26860) | 75% | 1456 | 10 | 9 | 3 | ATP-dependent serine protease | ||||
A0A0A0KW78 | Csa_4G017120 | TOM40-1 (AT3G20000) | 71% | 472 | 9 | 10 | 4 | Component of mitochondrial outer membrane translocase | ||||
A0A0A0LXK1 | Csa_1G629760 | SDH6 (AT1G08480) | 68% | 144 | 8 | 13 | 3 | Component of succinate dehydrogenase complex | ||||
A0A0A0KGU5 | Csa_6G500700 | MIC60 (AT4G39690) | 36% | 176 | 17 | 11 | 8 | Component of mitochondrial transmembrane lipoprotein complex | ||||
A0A0A0KGW6 | Csa_6G366300 | COS1 (AT2G44050) | 66% | 263 | 18,2 | 2,8E-06 | 5 | 6,7-Dimethyl-8-ribityllumazine synthase | ||||
A0A0A0KMM1 | Csa_5G047770 | AT1G14930 | 36% | 110 | 17 | Bet v1-type pathogenesis-related protein | ||||||
A0A0A0K9E8 | Csa_6G046410 | SD3 (AT4G00026) | 60% | 297 | 3 | 2 | 0 | Mitochondrial translocase | ||||
A0A0A0LSR2 | Csa_1G024260 | AT3G18240 | 62% | 506 | 13 | 9 | Mitochondrial ribosomal subunit | |||||
A0A0A0KKV4 | Csa_6G525450 | RFNR1 (AT4G05390) | 78% | 630 | 8 | 2 | Ferredoxin-NADP + reductase | |||||
A0A0A0LDA3 | Csa_3G435020 | MPPa1 (AT1G51980) | 63% | 616 | 7,4 | 1,2E-03 | 13 | 6 | 12 | Subunit alpha of mitochondrial processing peptidase complex | ||
A0A0A0L7Y6 | Csa_3G164480 | AT4G15940 | 76% | 347 | Fumaryl acetoacetate hydrolase | |||||||
A0A0A0KH76 | Csa_6G188090 | AT5G52370 | 57% | 151 | Mitochondrial 28S ribosomal protein S34 | |||||||
E1B2J6 | GAPDH | GAPC1 | 88% | 610 | 9 | Glyceraldehyde 3 phosphate dehydrogenase | ||||||
A0A0A0KBL8 | Csa_6G077460 | TKL2 (AT2G45290) | 83% | 1306 | 6,8 | 1,9E-03 | 0 | 5 | 5 | Transketolase | ||
A0A0A0KRJ5 | Csa_5G168830 | MKP11 (AT5G17165) | 51% | 89 | 9,7 | 2,4E-04 | Late embryogenesis abundant protein | |||||
A0A0A0M1R9_ A0A0A0LKR1 | Csa_1G574970 Csa_2G000830 | HXK1 (AT4G29130) | 74% | 757 | 11 | 4 | 3 | Hexokinase | ||||
A0A0A0KXY5 | Csa_4G337910 | AT5G63620 | 84% | 288 | 4 | 3 | 2 | Zinc-dependent alcohol dehydrogenase | ||||
A0A0A0LKD3 | Csa_2G346040 | UGP1 (AT5G17310) | 51% | 367 | 13 | 2 | 11 | UDP-glucose pyrophosphorylase | ||||
A0A0A0L2N5 | Csa_4G310720 | FAC1 (AT2G38280) | 80% | 1368 | 5,4 | 6,0E-03 | 11 | AMP deaminase | ||||
A0A0A0LZS4 | Csa_1G423090 | VDAC4 (AT5G57490) | 63% | 367 | 7 | 7 | 6 | Voltage-gated anion channel | ||||
A0A0A0LYA6 | Csa_1G532350 | AT4G33070 | 81% | 1042 | 3 | 6 | 5 | Thiamine pyrophosphate dependent pyruvate decarboxylase | ||||
A0A0A0LQ20 | Csa_2G403690 | CoxX3 (AT1G72020) | 66% | 138 | 16 | 12 | 9 | TonB-dependent heme receptor A | ||||
A0A0A0KGH2 | Csa_6G502730 | AT2G18330 | 75% | 891 | 2 | ATPase | ||||||
A0A0A0LLE7_ A0A0A0KLP6 | Csa_2G372170 Csa_6G497310 | SHM1 (AT4G37930) | 80% | 869 | 9 | 2 | 10 | Serine hydroxymethyl transferase | ||||
A0A0A0KGA1 | Csa_6G135470 | ALDH5F1 (AT1G79440) | 77% | 813 | 3,5 | 3,0E-02 | 7 | 3 | 9 | Succinate-semialdehyde dehydrogenase | ||
A0A0A0L5J6 | Csa_3G115030 | AT5G40810 | 87% | 514 | 3,4 | 3,6E-02 | 6 | 8 | 5 | Cytochrome c1 component of cyt-bc1 complex | ||
A0A0A0LP60 | Csa_2G360050 | SDH5 (AT1G47420) | 56% | 266 | 4 | 4 | 4 | Succinate dehydrogenase subunit 5 | ||||
A0A0A0KP30 | Csa_5G199270 | PIP1;4 (AT4G00430) | 87% | 523 | 15,4 | 1,0E-05 | 10 | 11 | 3 | Aquaporin | ||
A0A0A0LJB4 | Csa_2G010420 | ALDH2B7 (AT1G23800) | 80% | 888 | 18,3 | 2,7E-06 | 0 | 7 | 3 | Aldehyde dehydrogenase | ||
A0A0A0KYN6 | Csa_4G192110 | GDH1 (AT5G18170) | 91% | 788 | 24,1 | 2,7E-07 | 9 | 11 | 2 | Glutamate dehydrogenase | ||
A0A0A0KWX8 | Csa_4G050830 | PGD1 (AT1G64190) | 87% | 892 | 15,9 | 8,2E-06 | 3 | 4 | 1 | Phosphogluconate dehydrogenase | ||
A0A0A0KX20 | Csa_4G052590 | NDPK4 (AT4G23900) | 78% | 387 | 4,8 | 9,4E-03 | 15 | 2 | 1 | Nucleoside diphosphate kinase | ||
A0A0A0LHY6 | Csa_3G836500 | FDH (AT5G14780) | 83% | 628 | 15,4 | 1,0E-05 | 2 | 2 | 1 | Formate dehydrogenase | ||
A0A0A0K9Z6 | Csa_6G004600 | CYSC1 (AT3G61440) | 79% | 588 | 28,4 | 6,5E-08 | 8 | 11 | 0 | b-cyanoalanine synthase/cysteine synthase | ||
A0A0A0KKD9 | Csa_5G149330 | BIP2 (AT5G42020) | 91% | 1229 | 3,3 | 3,7E-02 | 14 | Heat shock protein 70 | ||||
A0A0A0K7B3 | Csa_7G222870 | CYS4 (AT4G16500) | 35% | 96 | 10,2 | 1,9E-04 | 5 | Cysteine-type endopeptidase inhibitor | ||||
A0A0A0LYF4 | Csa_1G710160 | Cysteine-type endopeptidase inhibitor | ||||||||||
A0A0A0KVK8 A0A0A0LRR3_ A0A0A0LDZ4 | Csa_4G094520 Csa_2G369070 Csa_3G889810 | GF14 (AT1G35160) | 93% | 482 | 14-3-3 protein | |||||||
A0A0A0K6A8 | Csa_7G070770 | TUA4 (AT1G04820) | 98% | 869 | 3,5 | 3,3E-02 | Tubulin |
Differential VDAC4 correlation degree in co-expression networks.
Formate dehydrogenase has been associated with stress responses in plants (
While FDH is co-expressed with a low number of proteins under Fe sufficiency, Fe starvation, and Mo sufficiency, such number strikingly increases under Mo starvation. Indeed, under this condition, FDH is co-expressed with 22 proteins (
ALDH2B7 and CYSC1 are themselves hubs under Mo starvation (
CYSC1 is co-expressed, under Mo starvation, with 16 proteins whereas it is co-expressed with 11 proteins, under Mo sufficiency (
AC4 interacts with tRNAs, and it might be involved in their transport into mitochondria (
VDAC4 is a co-expressed protein of the fumaryl acetoacetate hydrolase (
We hereby show how the topological network analysis applied to proteomes obtained from mitochondria purified from plants grown under Fe and/or Mo starvation suggests FDH as a hub of Mo nutrition in agreement with experimental observations (
Although the approaches based on computational prediction present some intrinsic limitations, including false-positive interactions or the lack of true ones, they nevertheless promote and support new experimental avenues, including large-scale experimental identification of PPIs, which will improve the effectiveness and accuracy of the proposed approaches.
The datasets presented in this study can be found in online repositories. The names of the repository/repositories and accession number(s) can be found in the article/
DD, PMo, GV, and IM conceived the work. DD reconstructed the PPI and co-expression networks with contributions from PMa and SH. IM, GV, and PMo analyzed all the networks and their biological relevance. IM wrote the manuscript with contributions from DD, GV, and PMo. All authors contributed to the article and approved the submitted version.
The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.
The Supplementary Material for this article can be found online at:
Homology between
Differentially expressed proteins and
Differential FDH correlation degree in co-expression networks.
Differential ALDH2B7 correlation degree in co-expression networks.
Differential CYSC1 correlation degree in co-expression networks.
Differential fumaryl acetoacetate hydrolase correlation degree in co-expression networks.
PPI hubs in the four nutritional conditions: control (+Fe+Mo), Mo starvation (+Fe−Mo), Fe starvation (−Fe+Mo), Mo and Fe starvation (−Fe−Mo).
PPI hubs in the four nutritional conditions: Fe sufficiency (consisting in +Mo and −Mo samples), Fe starvation (consisting in +Mo and −Mo samples), Mo sufficiency (consisting in +Fe and −Fe samples), and Mo starvation (consisting in +Fe and −Fe samples).
Co-expression and physical interaction under Fe sufficiency (+Fe+Mo and +Fe−Mo samples), Mo sufficiency (+Fe+Mo and −Fe+Mo), Fe starvation (−Fe+Mo, −Fe−Mo), and Mo starvation (+Fe−Mo, −Fe−Mo).