Comparative Dynamic Transcriptome Reveals the Delayed Secondary-Cell-Wall Thickening Results in Altered Lint Percentage and Fiber Elongation in a Chromosomal Segment Substitution Line of Cotton (Gossypium hirsutum L.)

Abstract

Lint percentage (LP) is an important yield component in cotton that is usually affected by initial fiber number and cell wall thickness. To explore how fiber cell wall development affects LP, phenotypic identification and dynamic transcriptome analysis were conducted using a single segment substitution line of chromosome 15 (SL15) that harbors a major quantitative trait locus (QTL) for LP. Compared to its recurrent parent LMY22, SL15 did not differ in initial fiber number, but the fiber cell wall thickness and single-fiber weight decreased significantly, altering LP. The comparative transcriptome profiles revealed that the secondary cell wall (SCW) development phase of SL15 was relatively delayed. Meanwhile, the expression of genes related to cell expansion decreased more slightly in SL15 with fiber development, resulting in relatively higher expression at SL15_25D than at LMY22_25D. SCW development-related genes, such as GhNACs and GhMYBs, in the putative NAC-MYB-CESA network differentially expressed at SL15_25D, along with the lower expression of CESA6, CSLC12, and CSLA2. The substituted chromosomal interval was further investigated, and found 6 of 146 candidate genes were differentially expressed in all four cell development periods including 10, 15, 20 and 25 DPA. Genetic variation and co-expression analysis showed that GH_D01G0052, GH_D01G0099, GH_D01G0100, and GH_D01G0140 may be important candidate genes associated with qLP-C15-1. Our results provide novel insights into cell wall development and its relationship with LP, which is beneficial for lint yield and fiber quality improvement.

Keywords: candidate genes; lint percentage; secondary-cell-wall thickening; single chromosomal segment substitution line; transcriptome analysis.

Figure 1

Multi-year field data comparisons. (A) Lint percentage and (B) fiber length in LMY22 and SL15 during a four-year field trial from 2016 to 2019 (mean ± s.d., n = 5, *P < 0.05, **P < 0.01, ***P < 0.001). (C) Representative fiber images from LMY22 (left) and SL15 (right). Scale bar, 3 cm.

Figure 2

Detection of fiber initiation and cell wall thickness. (A) TEM view of fiber initiation at 0 DPA; (i) and (ii) present the whole ovule of LMY22 and SL15, respectively, scale bar, 500 μm; (iii) and (iv) present close-up views of LMY22 and SL15, respectively, scale bar, 50 μm. (B) Counts of fiber initiations, mean ± s.d., n = 20. (C) Cross-sections of cotton fiber cells; (i) and (ii) present LMY22 and SL15 at 30 DPA, respectively; (iii) and (iv) present LMY22 and SL15 at 35 DPA, respectively; scale bar, 20 μm. (D) Ratio of cell wall thickness to cell radius (as percentage), mean ± s.d., n = 100, ***P < 0.001. (E) Weights of single fibers, mean ± s.d., n = 50, **P < 0.01 and ***P < 0.001.

Figure 3

Global transcriptome analysis. (A) Numbers of DEGs within and between LMY22 and SL15. (B) GO analysis comparing 20 DPA and 25 DPA in LMY22 and SL15.

Figure 4

Expression patterns of genes related to fiber elongation. Heat map for DEGs involved in (A) cell expansion and (B) cell wall loosening. (C) Expression of four DEGs as determined by qRT-PCR. *P < 0.05, **P < 0.01, and ***P < 0.001.

Figure 5

Expression patterns of genes in the NAC-MYB-CESA network. (A) Heat map for DEGs encoding NAC transcription factors. (B) Heat map for DEGs encoding MYB-like transcription factors and cellulose synthase/synthase-like genes. (C) Expression of six DEGs as determined by qRT-PCR. *P < 0.05, **P < 0.01, and ***P < 0.001.

Figure 6

Genetic structural variation and Expression patterns of candidate genes. (A) A schematic of introgressive segment on Chromosome 15 of SL15. (B) The genetic structural variation of candidate genes. Orange yellow boxes represent exons and black lines represent introns; the arrow represents the direction of genes on the chromosome. (C) Expression heat map for six candidate DEGs. (D) Expression of candidate genes as determined by qRT-PCR. *P < 0.05, **P < 0.01, and ***P < 0.001.

Figure 7

Model of fiber secondary cell wall thickening in LMY22 and SL15. Light gray and dark gray ovals respectively represent the cotton fiber primary cell wall and secondary cell wall (SCW); the orange oval represents the plasmalemma of the cotton fiber cell. Cylinders represent GhNACs that negatively regulate fiber SCW development, with orange and light gray indicating DEGs with up- or downregulated expression, respectively. Oblong waves represent MYBs involved in cellulose biosynthesis, with brick red and light green indicating DEGs with up- or downregulated expression, respectively. Hexagons represent cellulose synthase or cellulose-synthase-like genes, with bright red and light gray indicating DEGs with up- or downregulated expression, respectively. Colored dots represent various osmotically active solutes. Black arrows qualitatively represent the amount of water flowing from cytoplasm into vacuole.