Two distinct mechanisms silence chinmo in Drosophila neuroblasts and neuroepithelial cells to limit their self-renewal.

Whether common principles regulate the self-renewing potential of neural stem cells (NSCs) throughout the developing central nervous system is still unclear. In the Drosophila ventral nerve cord and central brain, asymmetrically dividing NSCs, called neuroblasts (NBs), progress through a series of sequentially expressed transcription factors that limits self-renewal by silencing a genetic module involving the transcription factor Chinmo. Here, we find that Chinmo also promotes neuroepithelium growth in the optic lobe during early larval stages by boosting symmetric self-renewing divisions while preventing differentiation. Neuroepithelium differentiation in late larvae requires the transcriptional silencing of chinmo by ecdysone, the main steroid hormone, therefore allowing coordination of neural stem cell self-renewal with organismal growth. In contrast, chinmo silencing in NBs is post-transcriptional and does not require ecdysone. Thus, during Drosophila development, humoral cues or tissue-intrinsic temporal specification programs respectively limit self-renewal in different types of neural progenitors through the transcriptional and post-transcriptional regulation of the same transcription factor.

Post-transcriptional control of light-harvesting genes expression under light stress.

Plants have to deal with fluctuating light environment and the regulation of the photosynthetic apparatus is crucial for their survival. The large multigenic family of nuclear encoded chloroplastic proteins called light harvesting complex (LHC) is involved in both light harvesting and photoprotection. Changes in light intensity induce a complex set of molecular events within both the chloroplast and the cytoplasmic compartments of the cell leading to reorganization of the photosynthetic apparatus in order to optimize photosynthesis to the new conditions. In this study we have investigated the occurrence of translational regulations during light stress in Arabidopsis thaliana by using polysomes profiling. We have observed a strong effect of light on global translation activity of the cell. We show that individual LHC genes are translationally regulated in response to light conditions by changing the ratio between polysomal versus total messenger RNA. In addition, we found that cytoplasmic translational regulation can precede nuclear transcriptional regulation. Thus translational control appears as an important component of the crosstalk between chloroplast and the nucleus in plant cells.

MYBL2 is a new regulator of flavonoid biosynthesis in Arabidopsis thaliana.

SUMMARY: In Arabidopsis thaliana, several MYB and basic helix-loop-helix (BHLH) proteins form ternary complexes with TTG1 (WD-Repeats) and regulate the transcription of genes involved in anthocyanin and proanthocyanidin (PA) biosynthesis. Similar MYB-BHLH-WDR (MBW) complexes control epidermal patterning and cell fates. A family of small MYB proteins (R3-MYB) has been shown to play an important role in the regulation of epidermal cell fates, acting as inhibitors of the MBW complexes. However, so far none of these small MYB proteins have been demonstrated to regulate flavonoid biosynthesis. The genetic and molecular analyses presented here demonstrated that Arabidopsis MYBL2, which encodes a R3-MYB-related protein, is involved in the regulation of flavonoid biosynthesis. The loss of MYBL2 activity in the seedlings of two independent T-DNA insertion mutants led to a dramatic increase in the accumulation of anthocyanin. In addition, overexpression of MYBL2 in seeds inhibited the biosynthesis of PAs. These changes in flavonoid content correlate well with the increased level of mRNA of several structural and regulatory anthocyanin biosynthesis genes. Interestingly, transient expression analyses in A. thaliana cells suggested that MYBL2 interacts with MBW complexes in planta and directly modulates the expression of flavonoid target genes. These results are fully consistent with the molecular interaction of MYBL2 with BHLH proteins observed in yeast. Finally, MYBL2 expression studies, including its inhibition by light-induced stress, allowed us to hypothesise a physiological role for MYBL2. Taken together, these results bring new insights into the transcriptional regulation of flavonoid biosynthesis and provide new clues and tools for further investigation of its developmental and environmental regulation.

Post-transcriptional regulation of gene expression in plants during abiotic stress.

Land plants are anchored in one place for most of their life cycle and therefore must constantly adapt their growth and metabolism to abiotic stresses such as light intensity, temperature and the availability of water and essential minerals. Thus, plants' subsistence depends on their ability to regulate rapidly gene expression in order to adapt their physiology to their environment. Recent studies indicate that post-transcriptional regulations of gene expression play an important role in how plants respond to abiotic stresses. We will review the different mechanisms of post-transcriptional regulation of nuclear genes expression including messenger RNA (mRNA) processing, stability, localization and protein translation, and discuss their relative importance for plant adaptation to abiotic stress.

Arabidopsis eIF2alpha kinase GCN2 is essential for growth in stress conditions and is activated by wounding.

BACKGROUND: Phosphorylation of eIF2alpha provides a key mechanism for down-regulating protein synthesis in response to nutrient starvation or stresses in mammalian and yeast cells. However, this process has not been well characterized in plants RESULTS: We show here that in response to amino acid and purine starvations, UV, cold shock and wounding, the Arabidopsis GCN2 kinase (AtGCN2) is activated and phosphorylates eIF2alpha. We show that AtGCN2 is essential for plant growth in stress situations and that its activity results in a strong reduction in global protein synthesis. CONCLUSION: Our results suggest that a general amino acid control response is conserved between yeast and plants but that the plant enzyme evolved to fulfill a more general function as an upstream sensor and regulator of diverse stress-response pathways. The activation of AtGCN2 following wounding or exposure to methyl jasmonate, the ethylene precursor 1-Aminocyclopropane-1-carboxylic acid (ACC) and salicylic acid, further suggests that this enzyme could play a role in plant defense against insect herbivores.

Detection of Argonaute 1 Association with Polysomes in Arabidopsis thaliana.

Argonaute (AGO) proteins play a key role in RNA silencing mechanisms. RNA silencing affects both RNA degradation and translation. The characterization of translation-associated RNA silencing mechanisms and components often requires polysome isolation and analysis. In this chapter, we describe the identification of AGO1 association with polysomes through polysome fractionation on sucrose gradient, preparation of proteins by filtration and concentration, and immunoblotting.

Neural stem cell-encoded temporal patterning delineates an early window of malignant susceptibility in Drosophila.

Pediatric neural tumors are often initiated during early development and can undergo very rapid transformation. However, the molecular basis of this early malignant susceptibility remains unknown. During Drosophila development, neural stem cells (NSCs) divide asymmetrically and generate intermediate progenitors that rapidly differentiate in neurons. Upon gene inactivation, these progeny can dedifferentiate and generate malignant tumors. Here, we find that intermediate progenitors are prone to malignancy only when born during an early window of development while expressing the transcription factor Chinmo, and the mRNA-binding proteins Imp/IGF2BP and Lin-28. These genes compose an oncogenic module that is coopted upon dedifferentiation of early-born intermediate progenitors to drive unlimited tumor growth. In late larvae, temporal transcription factor progression in NSCs silences the module, thereby limiting mitotic potential and terminating the window of malignant susceptibility. Thus, this study identifies the gene regulatory network that confers malignant potential to neural tumors with early developmental origins.

Building a brain under nutritional restriction: insights on sparing and plasticity from Drosophila studies.

While the growth of the developing brain is known to be well-protected compared to other organs in the face of nutrient restriction (NR), careful analysis has revealed a range of structural alterations and long-term neurological defects. Yet, despite intensive studies, little is known about the basic principles that govern brain development under nutrient deprivation. For over 20 years, Drosophila has proved to be a useful model for investigating how a functional nervous system develops from a restricted number of neural stem cells (NSCs). Recently, a few studies have started to uncover molecular mechanisms as well as region-specific adaptive strategies that preserve brain functionality and neuronal repertoire under NR, while modulating neuron numbers. Here, we review the developmental constraints that condition the response of the developing brain to NR. We then analyze the recent Drosophila work to highlight key principles that drive sparing and plasticity in different regions of the central nervous system (CNS). As simple animal models start to build a more integrated picture, understanding how the developing brain copes with NR could help in defining strategies to limit damage and improve brain recovery after birth.

Protection of neuronal diversity at the expense of neuronal numbers during nutrient restriction in the Drosophila visual system.

Systemic signals provided by nutrients and hormones are known to coordinate the growth and proliferation of different organs during development. However, within the brain, it is unclear how these signals influence neural progenitor divisions and neuronal diversity. Here, in the Drosophila visual system, we identify two developmental phases with different sensitivities to dietary nutrients. During early larval stages, nutrients regulate the size of the neural progenitor pool via insulin/PI3K/TOR-dependent symmetric neuroepithelial divisions. During late larval stages, neural proliferation becomes insensitive to dietary nutrients, and the steroid hormone ecdysone acts on Delta/Notch signaling to promote the switch from symmetric mitoses to asymmetric neurogenic divisions. This mechanism accounts for why sustained undernourishment during visual system development restricts neuronal numbers while protecting neuronal diversity. These studies reveal an adaptive mechanism that helps to retain a functional visual system over a range of different brain sizes in the face of suboptimal nutrition.

Biochemical evidence for translational repression by Arabidopsis microRNAs.

MicroRNAs (miRNAs) regulate gene expression posttranscriptionally through RNA silencing, a mechanism conserved in eukaryotes. Prevailing models entail most animal miRNAs affecting gene expression by blocking mRNA translation and most plant miRNAs, triggering mRNA cleavage. Here, using polysome fractionation in Arabidopsis thaliana, we found that a portion of mature miRNAs and ARGONAUTE1 (AGO1) is associated with polysomes, likely through their mRNA target. We observed enhanced accumulation of several distinct miRNA targets at both the mRNA and protein levels in an ago1 hypomorphic mutant. By contrast, translational repression, but not cleavage, persisted in transgenic plants expressing the slicing-inhibitor 2b protein from Cucumber mosaic virus. In agreement, we found that the polysome association of miR168 was lost in ago1 but maintained in 2b plants, indicating that translational repression is correlated with the presence of miRNAs and AGO1 in polysomes. This work provides direct biochemical evidence for a translational component in the plant miRNA pathway.