Multiple caleosins have overlapping functions in oil accumulation and embryo development

Abstract

Caleosins are lipid droplet and endoplasmic reticulum-associated proteins. To investigate their functions in oil accumulation, expression levels of caleosins in developing seeds of Arabidopsis thaliana were examined and four seed-expressed caleosins (CLO1, CLO2, CLO4 and CLO6) were identified. The four single mutants showed similar minor changes of fatty acid composition in seeds. Two double mutants (clo1 clo2 (CRISPR) and clo1×clo2 (crossing)) demonstrated distinct changes of fatty acid composition, 16-23% decrease of oil content and 10-13% decrease of seed weight. Moreover, a 40% decrease of oil content, further fatty acid changes and misshapen membrane of smaller lipid droplets were found in seeds of quadruple CLO RNAi lines. Notably, about 40% of quadruple CLO RNAi T1 seeds failed to germinate, and deformed embryos and seedlings were also found. Complementation experiments showed CLO1 rescued the phenotype of clo1 clo2. Overexpression of CLO1 in seedlings and BY2 cells increased triacylglycerol content up to 73.6%. Transcriptome analysis of clo1 clo2 developing seeds showed that expression levels of some genes related to lipid, embryo development, calcium signal and stress responses were affected. Taken together, our results suggest that the major seed-expressed caleosins have overlapping functions in oil accumulation and show pleiotropic effects on embryo development.

Caleosin 1 contributes to seed lipid droplet degradation by interaction with autophagy-related protein ATG8

“…20:1 level was also significantly lower in clo2 seeds. Similar changes in lipid composition in clo1 and clo2 were reported recently by Liu et al (2022) and linked to the function of both caleosins in lipid biosynthesis during A. thaliana seed development (Liu et al, 2022). Moreover, in silico analysis of Arabidopsis caleosins showed the presence of RY and RAV1 motifs in their promoters which can bind transcription factors regulating oil synthesis during seed development (Shen et al, 2014).…”

“…All these proteins contain an N-terminal hydrophilic domain consisting of an EF-hand calcium binding motif, a central hydrophobic region with the proline knot that anchors the protein to the LD membrane and a C-terminal hydrophilic region with several phosphorylation-and hemebinding sites (Purkrtova et al, 2007;Chapman et al, 2012;Huang, 2018). The genes encoding CLO1 and CLO2 are highly expressed in developing seeds suggesting their role in seed germination or dormancy (Poxleitner et al, 2006;Liu et al, 2022). Consequently, we examined here in detail the phenotypic effects of a CLO1 and CLO2 mutation on LD degradation during A. thaliana seed germination and provide evidence on the interaction of these proteins with ATG8.…”

Identification and Analysis of Expression Patterns of the Caleosin Genes in Hickory (Carya cathayensis Sarg.)

“…Liu discovered that doubly protruding seeds (clo1 × clo2) had a significant reduction in oil content and seed weight in Arabidopsis. In addition, overexpression of CLO1 in seedlings and BY2 cells increased triacylglycerol content up to 73.6% [35]. The expression of CcaCLO1 (Cca0849S0013) significantly increased during embryo development, indicating a potential response to oil accumulation in hickory.…”

A computational study on the structure–function relationships of plant caleosins

“…Reviewing other research met our results and partly opened up the knot of this puzzle. Accordingly, most CN and CNC caleosins had been reported as seed-specific proteins 6,9,[30][31][32][33][34][35][36][37][38][39][40][41][42][43][44] , while NC members had been isolated from the floral parts 9,17,42,[45][46][47][48][49][50][51] . That might explain the higher portion of NC and CNC in gymnosperms and angiosperms, respectively (Fig.…”