Genome-Wide Study of Plant-Specific PLATZ Transcription Factors and Functional Analysis of OsPLATZ1 in Regulating Caryopsis Development of Rice (Oryza sativa L.)

Abstract

Plant A/T-rich sequence- and zinc-binding protein (PLATZ) is a type of plant-specific zinc-dependent DNA-binding protein that binds to A/T-rich DNA sequences. This family is essential for plant growth, development, and stress response. In this study, 15 OsPLATZs were identified in the rice genome with complete PLATZ-conserved domains by CD-search, similar to those found in angiosperms. Multi-species phylogenetic analysis showed that PLATZs were conserved in photosynthetic organisms, and an evolutionary branch unique to angiosperms was identified among members of the PLATZ family. Fifteen OsPLATZs were represented by five groups, each with distinct characteristics. An analysis of protein structures and sequence motifs showed that OsPLATZs were similar within groups, but varied between them. The expression profile and qRT-PCR results showed that OsPLATZs had distinct expression patterns in different tissues, with some responding to stress induction. Most of the OsPLATZs localized to the nuclei, and were predicted to bind to DNA sequences by AlphaFold3, suggesting that they likely function as conventional transcription factors. We also identified OsPLATZ1, a caryopsis-specific gene that regulates grain filling and caryopsis development in rice. This research lays the foundation for exploring the structural diversity, evolutionary traits, expression profile, and possible roles of PLATZ transcription factors in rice.

Keywords: PLATZ; caryopsis development; expression pattern; photosynthetic organisms; rice.

Figure 1

Comparison of PLATZ distribution and phylogeny among sixteen photosynthetic organisms across evolutionary time. (A) Sixteen selected photosynthetic species representing a broad range of evolutionary time. (B) Phylogenetic tree of PLATZ was constructed to display the evolutionary history of PLATZ TF families.

Figure 2

Phylogenetic tree of PLATZ proteins in Oryza sativa, Zea mays, Arabidopsis thaliana, and Glycine max. The prefixes Os, Zm, At, and Gm before the gene name represent Oryza sativa, Zea mays, Arabidopsis thaliana, and Glycine max, respectively. The numbers under the branches refer to the bootstrap values of the maximum likelihood phylogenetic tree. The four groups are represented by different colors.

Figure 3

Collinearity analysis and mapping of OsPLATZs. (A) Collinear relationship analysis between orthologous PLATZ genes in rice, maize, Arabidopsis, and soybean. (B) Distribution and duplication of OsPLATZ genes in rice chromosomes. The tandem duplication event is marked in red. The chromosome numbers are shown at the top of each bar. The length of chromosomes was their relative extent. The scale on the left is in megabases (Mb).

Figure 4

Phylogenetic relationship, motif structure, and conservation analysis of OsPLATZs. (A) Maximum likelihood phylogenetic tree of OsPLATZ proteins. I-IV: OsPLATZs were divided into four groups and are represented using different colors; (B) MEME motif structure shows the distinct divergence between groups; (C) Batch-smart analysis of PLATZ domain distribution of OsPLATZ proteins.

Figure 5

The ratio of OsPLATZs whose promoters contain certain types of cis-acting elements. Cis-acting elements were identified by PlantCARE using the 2 kb fragment from upstream of the transcription start site of each OsPLATZ gene. The graph was generated based on the ratio of OsPLATZs (x-axis) whose promoters contain certain types of cis-acting elements related to different conditions (y-axis). The cis-acting elements related to cell cycle, transcription, development, hormone, abiotic/biotic stress, and polyadenylation machinery are represented by different colors.

Figure 6

Prediction of the tertiary structures of OsPLATZ proteins and their binding to DNA fragments. (A) Protein 3D structure prediction of OsPLATZs. Different colors distinguish individual members, and regions with low confidence are hidden. The dashed box represents the merged diagram of the “C”-shaped structures of the 15 rice PLATZs. (B) Prediction of binding between OsPLATZs protein and DNA fragment, where light blue represents α-helices, magenta represents β-strands, and metallic colors represent random coils. DNA fragments with A/T rich sequences published in pea were used to carry out protein–DNA docking analyses. Protein tertiary structure prediction based on AlphaFold3.

Figure 7

Expression patterns of OsPLATZ genes. (A) Expression patterns of OsPLATZs in different tissues analyzed using Plant Public RNA-seq data. The OsPLATZ genes are located on the right, and the tissues are at the bottom of each column. The color scale represents expression levels: red represents higher and white represents lower. The data values are shown as log₂ (FPKM). (B) qRT-PCR of OsPLATZs in different tissues and stages. R, root; S, stem; L, leaf; P, 3-6 cm panicle; O, ovary; A, anther; C, caryopsis; DAP, day after pollination.

Figure 8

Subcellular localization of OsPLATZs in rice protoplasts. pUbi::nGFP as control, H2B-mCherry fusion protein as a nuclear localization marker. GFP, green fluorescence of fusion proteins; mCherry, monomeric cherry fluorescent protein; Merged, merged microscopic images. Scale bars = 10 μm.

Figure 9

Phenotypic analyses of OsPLATZ1 knockout mutants. (A) Mature caryopses of ZH11, osplatz1-1, and osplatz1-2. Bars = 5 mm. (B) Surface views of cracked mature caryopses from ZH11, osplatz1-1, and osplatz1-2. Bars = 1 mm. (C) Chalky caryopses percentage of ZH11, osplatz1-1, and osplatz1-2 were quantified. n = 30, ** p < 0.01. (D) Statistical analysis of caryopses length, width, and thickness of ZH11, osplatz1-1, and osplatz1-2. n = 30, ** p < 0.01. ns, not significant. (E) Comparisons of 10-caryopses length, width, and thickness of ZH11, osplatz1-1, and osplatz1-2. Bars = 5 mm.