Transcript profiling of genes expressed during fibre development in diploid cotton (Gossypium arboreum L.)

Abstract

Background: Cotton fibre is a single cell and it is one of the best platforms for unraveling the genes express during various stages of fibre development. There are reports devoted to comparative transcriptome study on fiber cell initiation and elongation in tetraploid cultivated cotton. However, in the present investigation, comparative transcriptome study was made in diploid cultivated cotton using isogenic fuzzy-lintless (Fl) and normal fuzzy linted (FL) lines belong to Gossypium arboreum, diploid species at two stages, 0 and 10 dpa (days post anthesis), using Affymetrix cotton GeneChip genome array.

Result: Scanning electron microscopy (SEM) analysis uncovered the occurrence of few fibre cell initials in the Fl line as compared to many in Normal FL at -2 and 0 dpa. However, at 10 dpa there were no fibre cells found elongated in Fl but many elongated cells were found in FL line. Up-regulation of transcription factors, AP2-EREBP, C2H2, C3H, HB and WRKY was observed at 0 dpa whereas in 10 dpa transcription factors, AP2-EREBP, AUX/IAA, bHLH, C2H2, C3H, HB, MYB, NAC, Orphans, PLATZ and WRKY were found down regulated in Fl line. These transcription factors were mainly involved in metabolic pathways such as phytohormone signaling, energy metabolism of cell, fatty acid metabolism, secondary metabolism and other signaling pathways and are related directly or indirectly in fiber development. Quantitative real-time PCR was performed to check fold up or down-regulation of these genes and transcription factors (TFs) down regulated in mutants as compared to normal at 0 and 10 dpa.

Conclusion: This study elucidates that the up-regulation of transcription factors like AP2-EREBP, C2H2, C3H, HB, WRKY and phytohormone signaling genes at 0 dpa and their down-regulation at the 10 dpa might have constrain the fibre elongation in fuzzy-lintless line. Along with this the down-regulation of genes involved in synthesis of VLCFA chain, transcripts necessary for energy and cell wall metabolism, EXPANSINs, arabinogalactan proteins (AGPs), tubulin might also be the probable reason for reduced growth of fibres in the Fl. Plant receptor-like kinases (RLKs), Leucine Rich Repeats) LRR- family protein and signal transduction coding for mitogen-activated protein kinase (MAPK) cascade, have been engaged in coordination of cell elongation and SCW biosynthesis, down-regulation of these might loss the function leads to reduced fibre growth.

Keywords: Diploid cotton; Microarray; Transcription factors; qRT PCR.

Fig. 1

Images of developing fibre initials under Scanning Electron Microscopy (SEM). Images of SEM pictures of complete ovules (a, b, c, g, h, i) and epidermal layer of ovules showing development of fibre initials (d, e, f, j, k, l) in Gossypium arboreum (FL line) and its near isogenic FL line and mature seeds with elongated fibres in FL line (M) and in Fl line (N) are presented

Fig. 2

Transcriptome analysis during fibre development stages normal fuzzy linted (FL) lines and fuzzy-lintless (Fl) line. a Number of differentially expressed transcripts (DETs) in Fl line as compared to their respective stages in FL line at fibre initiation (0 dpa) and elongation (10 dpa) stages. b Venn diagram showing the commonly up- and down regulated transcripts between 0 and 10 dpa stages in Fl line. c Hierarchical cluster analysis of differentially expressed transcripts (fold change ≥3) in Fl line as compared to their respective stages in FL line at 0 dpa 10 dpa. A to E indicates the five major clusters. Cluster A: Commonly down regulated transcripts at 0 and 10 dpa in Fl line Cluster B: Only down regulated transcripts at 0 dpa in Fl line (8 DETs) Cluster C: Only up-regulated transcripts at 10 dpa in Fl line (15 DETs) Cluster D: Only up-regulated transcripts at 0 dpa in Fl line (42 DETs) Cluster E: Up-regulated at 0 dpa and down regulated at 10 dpa in Fl line (4 DETs) Cluster F: Only down regulated transcripts at 10 dpa in Fl line (197 DETs)

Fig. 3

Analysis of differently expressed transcripts (DETs) related to transcription factors (TFs) and phytohormone signaling (PHs) in Fl line compared to their respective stages in the FL line at fibre initation (0 dpa) and elongation (10 dpa) stages. a Number of DETs encoding TFs of various families in FL line at 0 and 10 dpa. b Number of DETs related to the phytohormone signaling in Fl line at 0 and 10 dpa

Fig. 4

Overview of differentially expressed transcripts present in various metabolic processes based on MapMan (version 3.5) visualization software in FL line at 0 dpa and 10 dpa 1: PS (Photo System), 2: Major CHO metabolism, 3: Minor CHO metabolism, 4: Glycolysis, 5: Fermentation, 6: Gluconeogensis/glycoxylate cycle, 7: OPP (O-phenylphenol), 8: TCA (tricarboxylic acid)/Org. transformation, 9: Mitochondrial electron transport/ATP synthesis, 10: Cell wall, 11: Lipid metabolism, 12: N-metabolism, 13: Amino acid metabolism, 14: S-assimilation, 15: Amino acid metabolism, 16: S-assimilation, 17: Hormone metabolism, 18: Co-factor and vitamine metabolism, 19: Tetrapyrrole synthesis, 20: Stress, 21: Redox, 22: Polyamine metabolism, 23: Nucletotide metabolism, 24: Biodegradation of Xenobiotics, 25: C1-metabolism, 26: Misc27: RNA, 28: DNA, 29: Protein, 29: Protein, 30: Signalling, 31: Cell, 32: Micro RNA, Natural antisense etc., 33: Development, 34: Transport, 35: Not assigned

Fig. 5

Validation of expression of genes obtained from microarray analysis in Fl line as compared to FL line at 0 dpa and 10 dpa