Biology Reference
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exhibiting higher levels of expression in the coa and spl mutants could be regarded
as candidate genes that were deregulated in the maternal sporophyte because of
the absence of a functional embryo sac in these mutants. Of the 527 genes identi-
fied for their maternal-gain-of-expression in coa and spl, about 9% were predicted
to be involved in transcriptional regulation and 7% were signaling proteins (Figure 1).
Among the genes encoding transcription factors, there were eight MYB class pro-
tein genes, seven zinc-finger protein genes including SUPERMAN and NUB-
BIN, five homeo box genes including SHOOT MERISTEMLESS (STM), five
genes each encoding basic helix-loop-helix (bHLH) and SQUAMOSA-binding
proteins, three genes encoding basic leucin zipper (bZIP) proteins, and two genes
each encoding APETALA2-domain and NAC-domain transcription factors. No
MADS box genes were represented. The genes encoding signaling proteins in-
cluded the auxin-responsive genes AUXIN RESISTANT 2/3 (AXR2 and AXR3),
three genes encoding DC1-domain-containing proteins, ten genes encoding ki-
nases and related proteins, two genes encoding phosphatases, four LRR-protein
genes, five auxin response regulator genes, and the two zinc-finger protein genes
SHORT INTERNODES (SHI) and STYLISH2 (STY2). When we examined
the whole dataset for genes encoding secreted proteins, 87 predicted proteins ful-
filled the criteria; 24% were below 20 kDa in size, which included a peptidase and
two lipid transfer proteins (data not shown).
The Carpel is the Major Target Tissue for Over-Expression
Caused by the Lack of an Embryo Sac
In order to confirm that the genes we identified truly reflect a gain of expression
in the maternal sporophyte of the mutant, we examined the expression levels and
patterns of 11 candidate genes in coa and wild-type gynoecium by RT-PCR or
in situ hybridization. Figure 6a shows an RT-PCR panel confirming that eight
genes from the coa dataset and three genes from the spl dataset were more highly
expressed in coa than in wild-type pistils. We present evidence that the genes we
identified for their gain of expression in spl were indeed over-expressed in coa as
well, suggesting that the genes are generally over-expressed in the absence of an
embryo sac, regardless of the mutation (Figure 6a). Figure 6 shows the expres-
sion of the following genes in the coa gynoecium as detected by in situ hybrid-
ization: AT4G12410 (a SAUR [auxin-responsive Small Auxin Up RNA] gene;
Figure 6b), AT1G75580 (an auxin-responsive gene; Figure 6c), AT5G03200 (en-
coding C3HC4-type RING finger protein; Figure 6d), AT5G15980 (encoding
PPR repeat-containing protein; Figure 6e), and STM (a homeo box gene; Figure
6g). Surprisingly, all of the five genes exhibited similar expression patterns: strong
expression in the carpel wall and septum, and relatively low expression in the
