A number of tissues are polarized production lines involved in the generation of highly specialized cell types. Examples include oogenesis within the gonad of many vertebrates and invertebrates and the crypt-villus axis of the mammalian gut (Simon and Gordon 1995
; Ko et al. 1997
). Oogenesis involves the constant production of oocytes (which are very large totipotent cells rich in cellular machinery, information molecules and nutrients) in a stepwise fashion for reproduction (Blumenfeld and Amit 1994; Matova and Cooley 2001). The generation of differentiated cells within such polarized tissue production-lines involves both developmental and physiological control mechanisms. Central to understanding polarized tissue function and homeostasis is uncovering the spatially integrated regulatory pathways that co-ordinate the developmental and physiological control of differentiated cell type production.
The germline of the adult C. elegans
hermaphrodite gonad is a polarized assembly line for the production of oocytes (; (Hirsh et al. 1976
)). In the presence of sperm, major sperm protein (MSP) induces oocyte maturation/ovulation every ~23 min (McCarter et al. 1999
; Miller et al. 2001
) achieving continuous oocyte production, a process that requires the biogenesis and organization of plasma membranes and cytoplasmic constituents, regulating gene expression as well as progression of chromosomes through meiotic prophase. Conversely, in middle-aged adult hermaphrodites (which have exhausted their sperm) or mutant females that lack male germ cells, oocytes already produced are arrested in late meiotic prophase and oocyte production is dramatically downregulated.
Summary of adult hermaphrodite germline development and the functions of the NOS-3/FEM-CUL-2/TRA-1 regulatory module
The RTK-RAS-ERK pathway relays physiological and developmental extracellular signals through a conserved kinase cascade that results in phosphorylation and activation of the extracellular-signal regulated kinase (ERK) (Sundaram et al. 1996
). Active ERK, in turn, controls biological processes through phosphorylation of substrate proteins (Chang and Karin 2001
). The C. elegans
ERK ortholog, MPK-1, controls at least seven different processes in hermaphrodite germline development, including membrane organization during oogenesis and progression of germ cell nuclei through pachytene of meiotic prophase (Lee et al. 2007
). Each of the seven processes is mediated by multiple MPK-1 substrates, with additional substrates likely remaining to be identified (Arur et al. 2009
). Activation of MPK-1 is (a) induced by the MSP signal and (b) spatially restricted to the medial and proximal regions of the oogenesis production line () where MPK-1 dependent processes are executed (Miller et al. 2001
; Lee et al. 2007
An essential prerequisite for oogenesis is the developmental specification of oocytes / female fate. In the hermaphrodite germline, the male fate (sperm) is specified during larval development and female fate (oocyte) is specified throughout adulthood. Germline sexual fate in C. elegans
is determined through an elaborate pathway involving more than 30 genes (Meyer 2005
; Zarkower 2006
; Ellis and Schedl 2007
), part of which is shown in . Important for this study is a multi-step negative regulatory module (NOS-3/FEM-CUL-2/TRA-1 module) necessary for oocyte fate specification. NOS-3, a homolog of the Nanos RNA binding protein, binds to FBF-1 and FBF-2 (FBF), two nearly identical Pumilio RNA binding protein homologs, which together repress translation of the fem-3
mRNA (Zhang et al. 1997
; Kraemer et al. 1999
). FEM-3 along with the sex determination proteins FEM-1 and FEM-2, forms a subunit of the CUL-2-based E3 ubiquitin ligase complex (Starostina et al. 2007
). As regulation of this CUL-2-based E3 ubiquitin ligase in the germline appears to be largely accomplished through FEM-3 levels and FEM-1, which is the substrate specificity subunit of the uniquitin ligase complex, we refer to it as the FEM-CUL-2 complex. The FEM-CUL-2 complex negatively regulates the TRA-1 Gli transcriptional repressor through ubiquitin mediated degradation (Starostina et al. 2007
). In the adult hermaphrodite, NOS-3 and FBF repress the translation of fem-3
mRNA resulting in the FEM-CUL-2 E3 ubiquitin ligase complex being inactive, leading to stabilization of nuclear TRA-1 Gli. For the specification of the oocyte fate, TRA-1 represses the transcription of fog-1
, genes that specify the sperm fate (Chen et al. 2000
; Jin et al. 2001
). Sexual fate is specified in the distal part of the germline, in the region of the germline stem cells (Crittenden et al. 2006
; Hansen and Schedl 2006
) determining the cell type generated within the gonadal production-line.
Here, we report an unexpected function of the NOS-3/FEM-CUL-2/TRA-1 module in regulation of plasma membrane organization during oogenesis. We demonstrate that MPK-1 promotes membrane organization by phosphorylation and inactivation of NOS-3, which through the negative regulatory module leads to degradation of nuclear TRA-1. We show that physiological control through MPK-1 and the NOS-3/FEM-CUL-2/TRA-1 module occurs in the medial and proximal germline where oocytes are being generated. By contrast, developmental control of sexual fate specification occurs in the distal germline. Thus, developmental and physiological mechanisms are integrated within the polarized germline tissue through spatially restricting the regulation and output of the NOS-3/FEM-CUL-2/TRA-1 module.