The novel plant protein INAPERTURATE POLLEN1 marks distinct cellular domains and controls formation of apertures in the Arabidopsis pollen exine.

Pollen grains protect the sperm cells inside them with the help of the unique cell wall, the exine, which exhibits enormous morphological variation across plant taxa, assembling into intricate and diverse species-specific patterns. How this complex extracellular structure is faithfully deposited at precise sites and acquires precise shape within a species is not understood. Here, we describe the isolation and characterization of the novel Arabidopsis thaliana gene INAPERTURATE POLLEN1 (INP1), which is specifically involved in formation of the pollen surface apertures, which arise by restriction of exine deposition at specific sites. Loss of INP1 leads to the loss of all three apertures in Arabidopsis pollen, and INP1 protein exhibits a unique tripartite localization in developing pollen, indicative of its direct involvement in specification of aperture positions. We also show that aperture length appears to be sensitive to INP1 dosage and INP1 misexpression can affect global exine patterning. Phenotypes of some inp1 mutants indicate that Arabidopsis apertures are initiated at three nonrandom positions around the pollen equator. The identification of INP1 opens up new avenues for studies of how formation of distinct cellular domains results in the production of different extracellular morphologies.

A large-scale genetic screen in Arabidopsis to identify genes involved in pollen exine production.

Exine, the outer plant pollen wall, has elaborate species-specific patterns, provides a protective barrier for male gametophytes, and serves as a mediator of strong and species-specific pollen-stigma adhesion. Exine is made of sporopollenin, a material remarkable for its strength, elasticity, and chemical durability. The chemical nature of sporopollenin, as well as the developmental mechanisms that govern its assembly into diverse patterns in different species, are poorly understood. Here, we describe a simple yet effective genetic screen in Arabidopsis (Arabidopsis thaliana) that was undertaken to advance our understanding of sporopollenin synthesis and exine assembly. This screen led to the recovery of mutants with a variety of defects in exine structure, including multiple mutants with novel phenotypes. Fifty-six mutants were selected for further characterization and are reported here. In 14 cases, we have mapped defects to specific genes, including four with previously demonstrated or suggested roles in exine development (MALE STERILITY2, CYP703A2, ANTHER-SPECIFIC PROTEIN6, TETRAKETIDE α-PYRONE REDUCTASE/DIHYDROFLAVONOL-4-REDUCTASE-LIKE1), and a number of genes that have not been implicated in exine production prior to this screen (among them, fatty acid ω-hydroxylase CYP704B1, putative glycosyl transferases At1g27600 and At1g33430, 4-coumarate-coenzyme A ligase 4CL3, polygalacturonase QUARTET3, novel gene At5g58100, and nucleotide-sugar transporter At5g65000). Our study illustrates that morphological screens of pollen can be extremely fruitful in identifying previously unknown exine genes and lays the foundation for biochemical, developmental, and evolutionary studies of exine production.