Germline stem cells are defined by their unique ability to generate more of themselves as well as differentiated gametes. The molecular mechanisms controlling the decision between self-renewal and differentiation are central unsolved problems in developmental biology with potentially broad medical implications. In Caenorhabditis elegans, germline stem cells are controlled by the somatic distal tip cell. FBF-1 and FBF-2, two nearly identical proteins, which together are called FBF ('fem-3 mRNA binding factor'), were originally discovered as regulators of germline sex determination. Here we report that FBF also controls germline stem cells: in an fbf-1 fbf-2 double mutant, germline proliferation is initially normal, but stem cells are not maintained. We suggest that FBF controls germline stem cells, at least in part, by repressing gld-1, which itself promotes commitment to the meiotic cell cycle. FBF belongs to the PUF family ('Pumilio and FBF') of RNA-binding proteins. Pumilio controls germline stem cells in Drosophila females, and, in lower eukaryotes, PUF proteins promote continued mitoses. We suggest that regulation by PUF proteins may be an ancient and widespread mechanism for control of stem cells.

FBF, a PUF RNA-binding protein, is a key regulator of the mitosis/meiosis decision in the Caenorhabditis elegans germline. Genetically, FBF has a dual role in this decision: it maintains germ cells in mitosis, but it also facilitates entry into meiosis. In this article, we explore the molecular basis of that dual role. Previous work showed that FBF downregulates gld-1 expression to promote mitosis and that the GLD-2 poly(A) polymerase upregulates gld-1 expression to reinforce the decision to enter meiosis. Here we ask whether FBF can act as both a negative regulator and a positive regulator of gld-1 expression and also investigate its molecular mechanisms of control. We first show that FBF co-immunoprecipitates with gld-1 mRNA, a result that complements previous evidence that FBF directly controls gld-1 mRNA. Then we show that FBF represses gld-1 expression, that FBF physically interacts with the CCF-1/Pop2p deadenylase and can stimulate deadenylation in vitro, and that CCF-1 is partially responsible for maintaining low GLD-1 in the mitotic region. Finally, we show that FBF can elevate gld-1 expression, that FBF physically interacts with the GLD-2 poly(A) polymerase, and that FBF can enhance GLD-2 poly(A) polymerase activity in vitro. We propose that FBF can affect polyadenylation either negatively by its CCF-1 interaction or positively by its GLD-2 interaction.

The Arabidopsis chy1 mutant is resistant to indole-3-butyric acid, a naturally occurring form of the plant hormone auxin. Because the mutant also has defects in peroxisomal ␤-oxidation, this resistance presumably results from a reduced conversion of indole-3-butyric acid to indole-3-acetic acid. We have cloned CHY1, which appears to encode a peroxisomal protein 43% identical to a mammalian valine catabolic enzyme that hydrolyzes ␤-hydroxyisobutyryl-CoA. We demonstrated that a human ␤-hydroxyisobutyryl-CoA hydrolase functionally complements chy1 when redirected from the mitochondria to the peroxisomes. We expressed CHY1 as a glutathione S-transferase (GST) fusion protein and demonstrated that purified GST-CHY1 hydrolyzes ␤-hydroxyisobutyryl-CoA. Mutagenesis studies showed that a glutamate that is catalytically essential in homologous enoyl-CoA hydratases was also essential in CHY1. Mutating a residue that is differentially conserved between hydrolases and hydratases established that this position is relevant to the catalytic distinction between the enzyme classes. It is likely that CHY1 acts in peroxisomal valine catabolism and that accumulation of a toxic intermediate, methacrylyl-CoA, causes the altered ␤-oxidation phenotypes of the chy1 mutant. Our results support the hypothesis that the energy-intensive sequence unique to valine catabolism, where an intermediate CoA ester is hydrolyzed and a new CoA ester is formed two steps later, avoids methacrylyl-CoA accumulation.

Although many genes that regulate floral development have been identified in Arabidopsis thaliana, relatively few are known in the grasses. In normal maize (Zea mays), each spikelet produces an upper and lower floral meristem, which initiate floral organs in a defined phyllotaxy before being consumed in the production of an ovule. The bearded-ear (bde) mutation affects floral development differently in the upper and lower meristem. The upper floral meristem initiates extra floral organs that are often mosaic or fused, while the lower floral meristem initiates additional floral meristems. We cloned bde by positional cloning and found that it encodes zea agamous3 (zag3), a MADS box transcription factor in the conserved AGAMOUS-LIKE6 clade. Mutants in the maize homolog of AGAMOUS, zag1, have a subset of bde floral defects. bde zag1 double mutants have a severe ear phenotype, not observed in either single mutant, in which floral meristems are converted to branch-like meristems, indicating that bde and zag1 redundantly promote floral meristem identity. In addition, BDE and ZAG1 physically interact. We propose a model in which BDE functions in at least three distinct complexes to regulate floral development in the maize ear.

RNA-binding proteins control germline development in metazoans. This work focuses on control of the C. elegans germline by two RNA-binding proteins: FOG-1, a CPEB homolog; and FBF, a PUF family member. Previous studies have shown that FOG-1 specifies the sperm fate and that FBF promotes proliferation. Here, we report that FOG-1 also promotes proliferation. Whereas fbf-1 fbf-2 double mutants make ∼120 germ cells, fog-1; fbf-1 fbf-2 triple mutants make only ∼10 germ cells. The triple mutant germline divides normally until early L2, when germ cells prematurely enter meiosis and begin oogenesis. Importantly, fog-1/+; fbf-1 fbf-2 animals make more germ cells than fbf-1 fbf-2 double mutants, demonstrating that one dose of wild-type fog-1promotes proliferation more effectively than two doses – at least in the absence of FBF. FOG-1 protein is barely detectable in proliferating germ cells, but abundant in germ cells destined for spermatogenesis. Based on fog-1 dose effects, together with the gradient of FOG-1 protein abundance, we suggest that low FOG-1 promotes proliferation and high FOG-1 specifies spermatogenesis. FBF binds specifically to regulatory elements in the fog-1 3′UTR, and FOG-1 increases in animals lacking FBF. Therefore, FBF represses fog-1 expression. We suggest that FBF promotes continued proliferation, at least in part, by maintaining FOG-1 at a low level appropriate for proliferation. The dose-dependent control of proliferation and cell fate by FOG-1 has striking parallels with Xenopus CPEB, suggesting a conserved mechanism in animal development.

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Inflorescence morphology is incredibly diverse. This diversity of form has been a fruitful source of inquiry for plant morphologists for more than a century. Work in the grasses (Poaceae), the tomato family (Solanaceae), and Arabidopsis thaliana (Brassicaceae) has led to a richer understanding of the molecular genetics underlying this diversity. The character of individual meristems, a combination of the number (determinacy) and nature (identity) of the products a meristem produces, is key in the development of plant form. A framework that describes inflorescence development in terms of shifting meristem identities has emerged and garnered empirical support in a number of model systems. We discuss this framework and highlight one important aspect of meristem identity that is often considered in isolation, phyllotaxis. Phyllotaxis refers to the arrangement of lateral organs around a central axis. The development and evolution of phyllotaxis in the inflorescence remains underexplored, but recent work analyzing early inflorescence development in the grasses identified an evolutionary shift in primary branch phyllotaxis in the Pooideae. We discuss the evidence for an intimate connection between meristem identity and phyllotaxis in both the inflorescence and vegetative shoot, and touch on what is known about the establishment of phyllotactic patterns in the meristem. Localized auxin maxima are instrumental in determining the position of lateral primordia. Upstream factors that regulate the position of these maxima remain unclear, and how phyllotactic patterns change over the course of a plant's lifetime and evolutionary time, is largely unknown. A more complete understanding of the molecular underpinnings of phyllotaxis and architectural diversity in inflorescences will require capitalizing on the extensive resources available in existing genetic systems, and developing new model systems that more fully represent the diversity of plant morphology.

Plant architecture is determined by meristems that initiate leaves during vegetative development and flowers during reproductive development. Maize (Zea mays) inflorescences are patterned by a series of branching events, culminating in floral meristems that produce sexual organs. The maize fuzzy tassel (fzt) mutant has striking inflorescence defects with indeterminate meristems, fasciation, and alterations in sex determination. fzt plants have dramatically reduced plant height and shorter, narrower leaves with leaf polarity and phase change defects. We positionally cloned fzt and discovered that it contains a mutation in a dicer-like1 homolog, a key enzyme required for microRNA (miRNA) biogenesis. miRNAs are small noncoding RNAs that reduce target mRNA levels and are key regulators of plant development and physiology. Small RNA sequencing analysis showed that most miRNAs are moderately reduced in fzt plants and a few miRNAs are dramatically reduced. Some aspects of the fzt phenotype can be explained by reduced levels of known miRNAs, including miRNAs that influence meristem determinacy, phase change, and leaf polarity. miRNAs responsible for other aspects of the fzt phenotype are unknown and likely to be those miRNAs most severely reduced in fzt mutants. The fzt mutation provides a tool to link specific miRNAs and targets to discrete phenotypes and developmental roles.