Regulation of gamete number and embryo sac cell identity
indeterminate gametophyte1 (ig1) has pleiotropic effects on development of the embryo sac and other plant parts. ig1 embryo sacs undergo extra rounds of free nuclear divisions resulting in extra eggs, extra central cells, and extra polar nuclei within central cells (Figure 1). Thus, wild-type ig1 function is most likely involved in limiting the number of free nuclear nuclear divisions. In the absence of ig1 function syncytial development is indeterminate.
Figure 1. Wild type (A) and ig1 (B) embryo sacs. The mutant embryo sac in B has three central cells with one nucleus each. The nuclei in two of the central cells are not located at the micropylar end as they would be in wild type.
The exact number and placement of nuclei at cellularization is variable in ig1 mutants. The phenotypes of ig1 embryo sacs suggest a position-based determination of cellular identity. The ability of the extra cells and nuclei to function as egg cells or polar nuclei, for example, may depend on their position in the embryo sac. Many of these defective embryo sacs give rise to abnormal seeds (Figure 2). Abnormalities include polyembryony, heterofertilization, miniature endosperms, and early abortion of seeds. The most obvious of these are the miniature and aborted seeds that occur in endosperms with a deviation from the normal 2 maternal : 1 paternal genome ratio—this occurs when there are extra polar nuclei in the central cell (Figure 2). Additionally ig1 restricts the embryogenic potential of cells that lack one of the two parental genomes. ig1 mutant embryo sacs produce haploid progeny, of both maternal and paternal origin, at a higher rate than wild type. The same seed phenotypes as in ig1 have been used to identify two more mutants with extra polar nuclei.
Figure 2. Ear of an ig1-O/ig1-O homozygous female pollinated by a homozygous wild type male. Many (but fewer than half) of the seeds are abnormal, including miniature and aborted kernels, which are easily recognized. The endosperm phenotype reflects the number of polar nuclei in the central cell before fertilization. Below the ear the relative sizes of seeds produced by fertilization of central cells with 2 polar nuclei (2 PN), 3 polar nuclei (3 PN), and 4 polar nuclei (4 PN).
The ig1 gene encodes a member of the LATERAL ORGANS BOUNDARIES (LOB) protein family with high similarity to ASYMMETRIC LEAVES2 (AS2) in Arabidopsis. Like as2 mutants, ig1 mutants have disrupted adaxial (upper) - abaxial (lower) leaf polarity with ectopic expression of abaxial leaf morphology (Figure 3).
Figure 3. Polarity defects in ig1 leaves. (Top left) Midrib of wild-type leaf. (Bottom left) Midrib and flaps of ig1 leaf. ig1 leaves have a patch of abaxial (lower surface) tissue on the upper surface of the leaf above the midrib and ectopic flaps of leaf blade that project from the upper surface on either side of the ectopic abaxial tissue. Arrowheads point to sclerenchyma cells, which are on the lower surface of wild-type leaves and both upper and lower surfaces of ig1 leaves. (Right) Model of adaxial and abaxial domains and leaf blade outgrowth in wild-type and ig1 leaves. The juxtaposition of abaxial and adaxial tissues produces a leaf margin, leading to blade outgrowth in wild type and blade and flap outgrowth in ig1. x=xylem. ph=phloem. cc=clear cells. Scale bar=150 µm.