High-throughput sequencing of partial cDNAs, or expressed sequence tags (ESTs), provides relatively fast and cost-effective access to the gene expression profile of an organism [1
]. EST libraries provide access to the population of genes transcribed, making analyses of ESTs informative in determining which genes are expressed at specific developmental ages, in specific tissues, or under specific environmental conditions.
EST analyses are especially useful when studying organisms for which little sequence data exists and for which sequencing of the genome is either not planned, or not easily feasible due to genome size. To date, there is little genomic data available for the Chlorophytes (green algae), a group far more diverse and evolutionarily divergent than all land plants combined. From this group, only Chlamydomonas reinhardtii
has been the object of an extensive EST project [3
]. Genomic information from this project proved critical to elucidating the function, biosynthesis, and regulation of the photosynthetic apparatus [4
(Fig. ), also known as the "Mermaid's Wineglass", is a giant unicellular green alga whose size and complex life cycle make it an attractive model system for understanding morphogenesis and subcellular localization [5
]. Reaching 3 cm in height at maturity, this unicell contains just a single diploid nucleus for most of its life cycle. It undergoes a complex morphogenetic program, most of which takes place at the apex [6
], centimeters away from the nucleus. Classic experiments on A. acetabulum
] provided the first compelling evidence for the role of the nucleus in morphogenesis and for the existence of "products of the nucleus", later presumed to be mRNAs [9
Figure 1 Juvenile, adult and reproductive morphologies of Acetabularia acetabulum. This giant alga has a complex life cycle and undergoes distinct developmental phases. From a spherical microscopic zygote, it initiates polarized growth elongating primarily at (more ...)
The life cycle of A. acetabulum
is composed of several developmental phases (Fig. ). Like multicellular land plants, juvenile and adult phases of A. acetabulum
are temporally sequential, but morphologically distinct [10
]. Juvenile phase comprises the first centimeter of growth while adult phase comprises the remaining 2 to 3 cm [10
]. Juvenile whorls of hairs are stacked closer to each other along the stalk, and the branching pattern of the hairs within each whorl is simpler than in adults [10
]. Physiologically, these two phases differ as well. For example, juveniles grow well in crowded conditions and poorly at low population densities, while adults grow well only at low population densities. Similar to land plants, the transition between phases is associated with a change in the reproductive competence of the apex [11
]. In A. acetabulum
, adult apices are competent to produce a terminal reproductive whorl, the cap, while juvenile apices are not (J Messmer and DF Mandoli, unpublished). At the molecular level however, the difference is gene expression patterns between adult and juvenile phases are virtually unknown.
To reveal differences in gene expression between adult and juvenile phases, we constructed two subtracted EST libraries from A. acetabulum
. These libraries were designed to contain transcripts specific to one phase or the other, presumably enriched in transcripts involved in morphogenesis or phase change. We randomly sequenced and analyzed 941 ESTs from these two libraries. Our analyses of these sequences indicate that juvenile and adult phases differ significantly in their gene expression patterns. We also identified 3 consensus sequences, shared mainly by adult ESTs, that have identity with introns and the 3'UTR from carbonic anhydrase genes we previously cloned [13
]. We discuss the potential role of these conserved elements in mRNA post-transcriptional regulation, particularly mRNA localization and/or stability.