No abstract

During the establishment of polarity, fucoid algal zygotes adhere to the substratum and select a growth axis according to environmental cues. Since little is known about the early events leading to axis selection, we investigated the chronology of cell adhesion, adhesive deposition, and axis selection induced by light (photopolarization). The requirements for secretion and the cytoskeleton in these processes and in the process of changing the orientation of an axis in response to new environmental cues (axis realignment) were also tested. Adhesive deposition occurred in two distinct stages: it was deposited uniformally on young zygotes (uniform primary adhesive) and later was deposited asymmetrically (polar secondary adhesive). Uniform primary adhesive deposition, cell adhesion, and photopolarization occurred simultaneously, and shortly thereafter, polar secondary adhesive deposition occurred at the future growth site. Uniform primary adhesive deposition and cell adhesion required secretion, but were independent of filamentous-actin (F-actin) and microtubule function. Photopolarization of young zygotes and polar secondary adhesive deposition required secretion but not microtubules. F-actin served to localize secondary adhesive deposition at the rhizoid pole; its function in polarization was more complex. F-actin was required for axis selection; however, its role in realignment of an axis depended on the light regime. The differing requirements for F-actin during development indicates that the axis is not static, but changes with time. These findings indicate that previous and future work on "axis formation" must be interpreted in the context of the developmental stage of the zygote.

Previous work has demonstrated that dynamic actin arrays are important for axis establishment and polar growth in the fucoid zygote, Silvetia compressa. Transitions between these arrays are mediated by depolymerization of an existing array and polymerization of a new array. To begin to understand how polymerization of new arrays might be regulated, we investigated the role of the highly conserved, actin-nucleating, Actin-related protein 2/3 (Arp2/3) complex. Arp2, a subunit of the complex, was cloned and peptide antibodies were raised to the C-terminal domain. In immunolocalization studies of polarizing zygotes, actin and Arp2 colocalized around the nucleus and in a patch at the rhizoid pole. In germinated zygotes, a cone of Arp2 and actin extended from the nucleus to the subapex. Within the rhizoid tip, three structural zones were observed in the majority of zygotes: the extreme apex was devoid of label, the subapex was enriched for Arp2, and further back both actin and Arp2 were present. This zonation suggests that actin nucleation occurs at the leading edge of the cone, in the Arp2-enriched region. In two sets of experiments, we showed that tip zonation is important for growth. First, pharmacological treatments that disrupted Arp2/actin zonation arrested tip growth. Second, changes in the direction of tip growth during negative phototropism were preceded by a reorientation of the zonation in accordance with the new growth direction. This work represents the first investigation of Arp2/3 complex localization in tip-growing algal cells.

Plants have evolved novel microtubule (MT) arrays to regulate cell division and cell expansion. How these MT arrangements are managed has been a question of long-standing interest to plant cell biologists. Do plants have unique ways of regulating MTs or have they co-opted mechanisms that are familiar to us from studies in animal and fungal cells? This Update focuses on the MT plus-end-tracking proteins (1TIPs), a relatively recent addition to the repertoire of MT regulatory proteins. Although the study of 1TIPs in plants is just beginning, the emerging data indicate that some, but not all, 1TIPs are conserved in the green lineage and that plants have at least one family of 1TIPs that is unique. Plants, it seems, organize their MT arrays via a combination of novel and phylogenetically conserved mechanisms.

The role of synthesis in the regulation of abscisic acid accumulation was investigated in the developing maize seed. To do this, expression and regulation of the abscisic acid biosynthetic enzyme phytoene desaturase were examined. Comparison of the gene sequence encoding phytoene desaturase and its transcript in the wild-type and viviparous-5 mutant showed that the mutant gene contains multiple insertions and deletions, resulting in the synthesis of a larger transcript. In addition, the 55-kDa phytoene desaturase protein was not detectable in the viviparous-5 mutant, indicating that this phenotype results from a mutation at the phytoene desaturase locus. Levels of phytoene desaturase transcript and protein were compared to abscisic acid levels during development to determine whether phytoene desaturase might regulate abscisic acid accumulation. In the endosperm, transcript levels were initially high and declined during late maturation and dormancy, while protein levels remained high throughout development. In the embryo, transcript levels were low and constant, while protein levels declined. Both temporal and tissue-specific expression of phytoene desaturase were unrelated to abscisic acid levels. An abscisic acid mutant (viviparous-2) deficient in phytoene desaturation was used to determine whether the wild-type protein encoded by Viviparous-2 regulates phytoene desaturase. Phytoene desaturase transcript and protein levels were compared in wild-type and viviparous-2 mutant embryos and endosperm. Normalized levels of phytoene desaturase were similar in wild-type and mutant tissues, suggesting that the wild-type Viviparous-2 protein does not regulate phytoene desaturase transcript or protein levels.