CAND1 is required for pollen viability in Arabidopsis thaliana—a test of the adaptive exchange hypothesis

Frontiers in Plant Science · 2022-07-28 · 6 citations

The dynamic assembly of SKP1•CUL1•F-box protein (SCF) ubiquitin ligases is important for protein ubiquitination and degradation. This process is enabled by CAND1, which exchanges F-box proteins associated with the common CUL1 scaffold, and thereby, recycles the limited CUL1 core and allows diverse F-box proteins to assemble active SCFs. Previous human cell biological and computational studies have led to the adaptive exchange hypothesis, which suggests that the CAND1-mediated exchange confers plasticity on the SCF system, allowing cells to tolerate large variations in F-box protein expression. Here, we tested this hypothesis using Arabidopsis thaliana , a multicellular organism expressing hundreds of F-box protein genes at variable levels in different tissues. The cand1 null mutant in Arabidopsis is viable but produce almost no seeds. Bioinformatic, cell biological, and developmental analyses revealed that the low fertility in the cand1 mutant is associated with cell death in pollen, where the net expression of F-box protein genes is significantly higher than any other Arabidopsis tissue. In addition, we show that the transmission efficiency of the cand1 null allele was reduced through the male but not the female gametophyte. Our results suggest that CAND1 activity is essential in cells or tissues expressing high levels of F-box proteins. This finding is consistent with the proposed adaptive exchange hypothesis, demonstrating the necessity of the evolutionarily conserved CAND1-mediated exchange system in the development of a multicellular organism.

AtNOT1 is required for gametophyte development in Arabidopsis

The Plant Journal · 2020 · 18 citations

• Biology
• Genetics
• Cell biology

In flowering plants, pollen development is under a dynamic and well-orchestrated transcriptional control, characterized by an early phase with high transcript diversity and a late post-mitotic phase skewed to a cell-type-specific transcriptome. Such transcriptional changes require a balance between synthesis and degradation of mRNA transcripts, the latter being initiated by deadenylation. The CCR4-NOT complex is the main evolutionary conserved deadenylase complex in eukaryotes, and its function is essential during germline specification in animals. We hypothesized that the CCR4-NOT complex might play a central role in mRNA turnover during microgametogenesis in Arabidopsis. Disruption of NOT1 gene, which encodes the scaffold protein of the CCR4-NOT complex, showed abnormal seed set. Genetic analysis failed to recover homozygous progeny, and reciprocal crosses confirmed reduced transmission through the male and female gametophytes. Concordantly, not1 embryo sacs showed delayed development and defects in embryogenesis. not1 pollen grains exhibited abnormal male germ unit configurations and failed to germinate. Transcriptome analysis of pollen from not1/+ mutants revealed that lack of NOT1 leads to an extensive transcriptional deregulation during microgametogenesis. Therefore, our work establishes NOT1 as an important player during gametophyte development in Arabidopsis.

Targeted reprogramming of H3K27me3 resets epigenetic memory in plant paternal chromatin

Nature Cell Biology · 2020 · 228 citations

• Biology
• Cell biology
• Genetics

Plant Cell Wall Composition: Does Ploidy Matter?

PLANT PHYSIOLOGY · 2019-01-01 · 3 citations

Most of the carbon dioxide sequestered by plants during photosynthesis is converted into sugars and stored into polysaccharide-enriched cells walls that constitute the majority of the plant biomass. While plants have long been considered a valuable resource of biomaterials for the chemical and

Live-Cell Imaging of Mobile RNAs in Plants

PLANT PHYSIOLOGY · 2018-06-01

One of the most exciting findings in the past few decades is the discovery that individual mRNAs and noncoding RNAs can act as long-distance signaling messengers traveling cell to cell to distant sites in the plant. Numerous examples unveiled the involvement of endogenous RNAs as non-cell-autonomous

Plant Evolution: What Does It Take To Be an Egg?

Current Biology · 2016-07-01 · 2 citations

Intercellular communication in <i>Arabidopsis thaliana</i> pollen discovered via <i>AHG3</i> transcript movement from the vegetative cell to sperm

Proceedings of the National Academy of Sciences · 2015-10-14 · 27 citations

An Arabidopsis pollen grain (male gametophyte) consists of three cells: the vegetative cell, which forms the pollen tube, and two sperm cells enclosed within the vegetative cell. It is still unclear if there is intercellular communication between the vegetative cell and the sperm cells. Here we show that ABA-hypersensitive germination3 (AHG3), encoding a protein phosphatase, is specifically transcribed in the vegetative cell but predominantly translated in sperm cells. We used a series of deletion constructs and promoter exchanges to document transport of AHG3 transcripts from the vegetative cell to sperm and showed that their transport requires sequences in both the 5' UTR and the coding region. Thus, in addition its known role in transporting sperm during pollen tube growth, the vegetative cell also contributes transcripts to the sperm cells.

Setting the Stage for the Next Generation: Epigenetic Reprogramming During Sexual Plant Reproduction

2015-01-01 · 1 citations

Arabidopsis Tetraspanins Are Confined to Discrete Expression Domains and Cell Types in Reproductive Tissues and Form Homo- and Heterodimers When Expressed in Yeast      

PLANT PHYSIOLOGY · 2013-08-15 · 88 citations

Tetraspanins are evolutionary conserved transmembrane proteins present in all multicellular organisms. In animals, they are known to act as central organizers of membrane complexes and thought to facilitate diverse biological processes, such as cell proliferation, movement, adhesion, and fusion. The genome of Arabidopsis (Arabidopsis thaliana) encodes 17 members of the tetraspanin family; however, little is known about their functions in plant development. Here, we analyzed their phylogeny, protein topology, and domain structure and surveyed their expression and localization patterns in reproductive tissues. We show that, despite their low sequence identity with metazoan tetraspanins, plant tetraspanins display the typical structural topology and most signature features of tetraspanins in other multicellular organisms. Arabidopsis tetraspanins are expressed in diverse tissue domains or cell types in reproductive tissues, and some accumulate at the highest levels in response to pollination in the transmitting tract and stigma, male and female gametophytes and gametes. Arabidopsis tetraspanins are preferentially targeted to the plasma membrane, and they variously associate with specialized membrane domains, in a polarized fashion, to intercellular contacts or plasmodesmata. A membrane-based yeast (Saccharomyces cerevisiae) two-hybrid system established that tetraspanins can physically interact, forming homo- and heterodimer complexes. These results, together with a likely genetic redundancy, suggest that, similar to their metazoan counterparts, plant tetraspanins might be involved in facilitating intercellular communication, whose functions might be determined by the composition of tetraspanin complexes and their binding partners at the cell surface of specific cell types.

FACS-based purification of Arabidopsis microspores, sperm cells and vegetative nuclei

Plant Methods · 2012-10-17 · 94 citations

BACKGROUND: The male germline in flowering plants differentiates by asymmetric division of haploid uninucleated microspores, giving rise to a vegetative cell enclosing a smaller generative cell, which eventually undergoes a second mitosis to originate two sperm cells. The vegetative cell and the sperm cells activate distinct genetic and epigenetic mechanisms to control pollen tube growth and germ cell specification, respectively. Therefore, a comprehensive characterization of these processes relies on efficient methods to isolate each of the different cell types throughout male gametogenesis. RESULTS: We developed stable transgenic Arabidopsis lines and reliable purification tools based on Fluorescence-Activated Cell Sorting (FACS) in order to isolate highly pure and viable fractions of each cell/nuclei type before and after pollen mitosis. In the case of mature pollen, this was accomplished by expressing GFP and RFP in the sperm and vegetative nuclei, respectively, resulting in 99% pure sorted populations. Microspores were also purified by FACS taking advantage of their characteristic small size and autofluorescent properties, and were confirmed to be 98% pure. CONCLUSIONS: We provide simple and efficient FACS-based purification protocols for Arabidopsis microspores, vegetative nuclei and sperm cells. This paves the way for subsequent molecular analysis such as transcriptomics, DNA methylation analysis and chromatin immunoprecipitation, in the developmental context of microgametogenesis in Arabidopsis.