Judith Kimble

Asymmetric and symmetric stem-cell divisions in development and cancer

Listen to an interview with Sean Morrison on the stem cells podcastMuch has been made of the idea that asymmetric cell division is a defining characteristic of stem cells that enables them to simultaneously perpetuate themselves (self-renew) and generate differentiated progeny. Yet many stem cells can divide symmetrically, particularly when they are expanding in number during development or after injury. Thus, asymmetric division is not necessary for stem-cell identity but rather is a tool that stem cells can use to maintain appropriate numbers of progeny. The facultative use of symmetric or asymmetric divisions by stem cells may be a key adaptation that is crucial for adult regenerative capacity.

glp-1 Is required in the germ line for regulation of the decision between mitosis and meiosis in C. elegans

In the wild-type C. elegans germ line there are both mitotic and meiotic germ cells. Mutations in glp-1 cause germ cells that would normally divide mitotically to enter meiosis. This mutant phenotype mimics the effect of killing the distal tip cell, a somatic cell that interacts with the germ line to regulate the mitotic/meiotic decision. In addition, wild-type glp-1 product is required maternally for embryogenesis. Temperature-shift experiments indicate that the temporal requirement for glp-1 activity in the germ line is the same as that for distal tip cell regulation. Mosaic analyses suggest that glp-1 is produced in the germ line. We propose that glp-1 acts as part of the receiving mechanism in the interaction between the distal tip cell and germ line.

On the control of germ cell development in Caenorhabditis elegans

Abstract After hatching, the germ line progenitor cells in C. elegans begin to divide mitotically; later, some of the germ line cells enter meiosis and differentiate into gametes. In the adult, mitotic germ cells, or stem cells, are found at one end (the distal end) and meiotic cells occupy the rest of the elongate gonad. Removal of two somatic gonadal cells, the distal tip cells, by laser microsurgery has a dramatic effect on germ cell development. In either sex, this operation leads to the arrest of mitosis and the initiation of meiosis in germ cells. The function of the distal tip cell in the intact animal appears to be the inhibition of meiosis (or stimulation of mitosis) in nearby germ cells. During development, this permits growth and, in the adult, it maintains the germ line stem cell population. A change in the position of the distal tip cell in the gonad at an early point in development is correlated with a change in the axial polarity of the germ line tissue. This suggests that the localization of the distal tip cells inhibitory activity at the distal end of the gonad establishes the axial polarity of the germ line tissue in the intact animal.

A conserved RNA-binding protein that regulates sexual fates in the C. elegans hermaphrodite germ line

The nematode Caenorhabditis elegans has two sexes, males and hermaphrodites. Hermaphrodites initially produce sperm but switch to producing oocytes. This switch appears to be controlled by the 3′ untranslated region of fem-3 messenger RNA. We have now identified a binding factor (FBF) which is a cytoplasmic protein that binds specifically to the regulatory region of fem-3 3′UTR and mediates the sperm/oocyte switch. The RNA-binding domain of FBF consists of a stretch of eight tandem repeats and two short flanking regions. This structural element is conserved in several proteins including Drosophila Pumilio, a regulatory protein that controls pattern formation in the fly by binding to a 3′UTR. We propose that FBF and Pumilio are members of a widespread family of sequence-specific RNA-binding proteins.

Alterations in cell lineage following laser ablation of cells in the somatic gonad of Caenorhabditis elegans

Abstract The postembryonic cell lineage of the somatic gonad is essentially invariant in Caenorhabditis elegans ( J.E. Kimble and D. Hirsh, 1979, Develop. Biol. 70, 396–417 ). The two exceptions to this rule of invariance involve a natural ambiguity in the ancestry of certain cells such that each of two precursor cells assumes one of two alternative fates in a given animal. In this paper, experiments are reported in which laser microsurgery is used to kill individual cells in the developing somatic gonad. Such intervention perturbs the normal environment of the remaining cells; a change observed in the expected behavior of these cells suggests that extrinsic cues may normally play a role in controlling that behavior. Several different lineage alterations have been observed after laser microsurgery in the somatic gonad. These include switches in the type of lineage followed by a given precursor cell, reversals in lineage polarity, duplications of a lineage, and alteratiions in the number of cells produced in the lineage. The only cases in which cells switch from one lineage type to another involve pairs of cells which exhibit natural ambiguity. In most cases, the interactions inferred from these changes seem to occur between neighboring somatic gonadal cells. In one case, induction of the vulva, the interaction occurs between a single somatic gonadal cell, the anchor cell, and the precursors to the vulva in a neighboring tissue, the hypodermis. The roles of intrinsic and extrinsic cues in controlling normally invariant cell lineages are discussed.

A conserved RNA-binding protein controls germline stem cells in Caenorhabditis elegans

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.

A regulatory cytoplasmic poly(A) polymerase in Caenorhabditis elegans.

Messenger RNA regulation is a critical mode of controlling gene expression. Regulation of mRNA stability and translation is linked to controls of poly(A) tail length. Poly(A) lengthening can stabilize and translationally activate mRNAs, whereas poly(A) removal can trigger degradation and translational repression. Germline granules (for example, polar granules in flies, P granules in worms) are ribonucleoprotein particles implicated in translational control. Here we report that the Caenorhabditis elegans gene gld-2, a regulator of mitosis/meiosis decision and other germline events, encodes the catalytic moiety of a cytoplasmic poly(A) polymerase (PAP) that is associated with P granules in early embryos. Importantly, the GLD-2 protein sequence has diverged substantially from that of conventional eukaryotic PAPs, and lacks a recognizable RRM (RNA recognition motif)-like domain. GLD-2 has little PAP activity on its own, but is stimulated in vitro by GLD-3. GLD-3 is also a developmental regulator, and belongs to the Bicaudal-C family of RNA binding proteins. We suggest that GLD-2 is the prototype for a class of regulatory cytoplasmic PAPs that are recruited to specific mRNAs by a binding partner, thereby targeting those mRNAs for polyadenylation and increased expression.

Tissue-specific synthesis of yolk proteins in Caenorhabditis elegans☆

The primary site of yolk protein synthesis in the nematode, Caenorhabditis elegans, has been determined. In animals containing no gonadal cells (obtained by laser ablation of the gonadal precursor cells early in development), yolk proteins are present in abundance. This demonstrates that yolk proteins are made outside the gonad. An examination of proteins present in tissues isolated by dissection, and a comparison of proteins synthesized by isolated tissues incubated in vitro have identified the intestine as the major site of yolk protein synthesis. We propose that yolk proteins are synthesized in the intestine, secreted from the intestine into the body cavity, and taken up from the body cavity by the gonad to reach oocytes. The site of yolk protein synthesis has also been examined in four mutants that have largely male somatic tissues, but a hermaphrodite germ line. Here again, yolk proteins are produced by intestines in a hermaphrodite-specific manner. This suggests that sex determination is coordinately regulated in intestinal and germ line tissues.

A nuclear Argonaute promotes multigenerational epigenetic inheritance and germline immortality

Epigenetic information is frequently erased near the start of each new generation. In some cases, however, epigenetic information can be transmitted from parent to progeny (multigenerational epigenetic inheritance). A particularly notable example of this type of epigenetic inheritance is double-stranded RNA-mediated gene silencing in Caenorhabditis elegans. This RNA-mediated interference (RNAi) can be inherited for more than five generations. To understand this process, here we conduct a genetic screen for nematodes defective in transmitting RNAi silencing signals to future generations. This screen identified the heritable RNAi defective 1 (hrde-1) gene. hrde-1 encodes an Argonaute protein that associates with small interfering RNAs in the germ cells of progeny of animals exposed to double-stranded RNA. In the nuclei of these germ cells, HRDE-1 engages the nuclear RNAi defective pathway to direct the trimethylation of histone H3 at Lys 9 (H3K9me3) at RNAi-targeted genomic loci and promote RNAi inheritance. Under normal growth conditions, HRDE-1 associates with endogenously expressed short interfering RNAs, which direct nuclear gene silencing in germ cells. In hrde-1- or nuclear RNAi-deficient animals, germline silencing is lost over generational time. Concurrently, these animals exhibit steadily worsening defects in gamete formation and function that ultimately lead to sterility. These results establish that the Argonaute protein HRDE-1 directs gene-silencing events in germ-cell nuclei that drive multigenerational RNAi inheritance and promote immortality of the germ-cell lineage. We propose that C. elegans use the RNAi inheritance machinery to transmit epigenetic information, accrued by past generations, into future generations to regulate important biological processes.

Translational control of maternal glp-1 mRNA establishes an asymmetry in the C. elegans embryo

Abstract In C. elegans, the glp-1 gene encodes a membrane receptor that is required for anterior cell fates in the early embryo. We report that GLP-1 protein is localized to anterior blastomeres in 2- to 28-cell embryos. By contrast, glp-1 mRNA is present in all blastomeres until the 8-cell stage. Furthermore, the glp-1 3′ untranslated region can restrict translation of a reporter mRNA to anterior blastomeres. Therefore, the translation of maternal glp-1 mRNA is temporally and spatially regulated in the C. elegans embryo. The regulation of maternal glp-1 mRNA has striking parallels to the regulation of maternal hunchback mRNA in the Drosophila embryo. Thus, the establishment of embryonic asymmetry in diverse organisms may involve conserved mechanisms of maternal mRNA regulation.