BIBLIOGRAPHY

1. Maciejowski, John, Ugel, Nadia, Mishra, Bud, Isopi, Marco, Hubbard, E Jane Albert. "Quantitative analysis of germline mitosis in adult C. elegans," Developmental biology (Orlando) 2006 Apr 1;292(1):142-51. Epub 2006 Feb 15.   (MEDL:16480707 PMID: 16480707 #J0125773)    

   Certain aspects of the distal gonad of C. elegans are comparable to niche/stem cell systems in other organisms. The distal tip cell (DTC) caps a blind-ended tube; only the distal germ cells maintain proliferation in response to signaling from the DTC via the GLP-1/Notch signaling pathway in the germ line. Fruitful comparison between this system and other stem cell systems is limited by a lack of basic information regarding germ cell division behavior in C. elegans. Here, we explore the spatial pattern of cell division frequency in the adult C. elegans germ line relative to distance from the distal tip. We mapped the positions of actively dividing germline nuclei in over 600 fixed gonad preparations including the wild type and a gain-of-function ligand-responsive GLP-1 receptor mutant with an extended mitotic zone. One particularly surprising observation from these data is that the frequency of cell divisions is lower in distal-most cells-cells that directly contact the distal tip cell body-relative to cells further proximal, a difference that persists in the gain-of-function GLP-1 mutant. These results suggest that cell division frequency in the distal-most cells may be suppressed or otherwise controlled in a complex manner. Further, our data suggest that the presence of an active cell division influences the probability of observing simultaneous cell divisions in the same gonad arm, and that simultaneous divisions tend to cluster spatially. We speculate that this system behaves similarly to niche/stem cell/transit amplifying cell systems in other organisms.

2. Voutev, Roumen, Killian, Darrell J, Ahn, James Hyungsoo, Hubbard, E Jane Albert. "Alterations in ribosome biogenesis cause specific defects in C. elegans hermaphrodite gonadogenesis," Developmental biology (Orlando) 2006 Oct 1;298(1):45-58. Epub 2006 Jun 8.   (MEDL:16876152 PMID: 16876152 #J0126775)    

   Ribosome biogenesis is a cell-essential process that influences cell growth, proliferation, and differentiation. How ribosome biogenesis impacts development, however, is poorly understood. Here, we establish a link between ribosome biogenesis and gonadogenesis in Caenorhabditis elegans that affects germline proliferation and patterning. Previously, we determined that pro-1(+)activity is required in the soma--specifically, the sheath/spermatheca sublineage--to promote normal proliferation and prevent germline tumor formation. Here, we report that PRO-1, like its yeast ortholog IPI3, influences rRNA processing. pro-1 tumors are suppressed by mutations in ncl-1 or lin-35/Rb, both of which elevate pre-rRNA levels. Thus, in this context, lin-35/Rb acts as a soma-autonomous germline tumor promoter. We further report the characterization of two additional genes identified for their germline tumor phenotype, pro-2 and pro-3, and find that they, too, encode orthologs of proteins involved in ribosome biogenesis in yeast (NOC2 and SDA1, respectively). Finally, we demonstrate that depletion of additional C. elegans orthologs of yeast ribosome biogenesis factors display phenotypes similar to depletion of progenes. We conclude that the C. elegans distal sheath is particularly sensitive to alterations in ribosome biogenesis and that ribosome biogenesis defects in one tissue can non-autonomously influence proliferation in an adjacent tissue.

3. Fisher, Jasmin, Piterman, Nir, Hubbard, E Jane Albert, Stern, Michael J, Harel, David. "Computational insights into Caenorhabditis elegans vulval development," Proceedings of the National Academy of Sciences of the United States of America 2005 Feb 8;102(6):1951-6. Epub 2005 Jan 31.   (MEDL:15684055 PMID: 15684055 #J0126778)    

   Studies of Caenorhabditis elegans vulval development provide a paradigm for pattern formation during animal development. The fates of the six vulval precursor cells are specified by the combined action of an inductive signal that activates the EGF receptor mitogen-activated PK signaling pathway (specifying a primary fate) and a lateral signal mediated by LIN-12/Notch (specifying a secondary fate). Here we use methods devised for the engineering of complex reactive systems to model a biological system. We have chosen the visual formalism of statecharts and use it to formalize Sternberg and Horvitz's 1989 model [Sternberg, P. W. & Horvitz, H. R. (1989) Cell 58, 679-693], which forms the basis for our current understanding of the interaction between these two signaling pathways. The construction and execution of our model suggest that different levels of the inductive signal induce a temporally graded response of the EGF receptor mitogen-activated PK pathway and make explicit the importance of this temporal response. Our model also suggests the existence of an additional mechanism operating during lateral specification that prohibits neighboring vulval precursor cells from assuming the primary fate.

4. Killian, Darrell J, Hubbard, E Jane Albert. "Caenorhabditis elegans germline patterning requires coordinated development of the somatic gonadal sheath and the germ line," Developmental biology (Orlando) 2005 Mar 15;279(2):322-35.   (MEDL:15733661 PMID: 15733661 #J0126776)    

   Interactions between the somatic gonad and the germ line influence the amplification, maintenance, and differentiation of germ cells. In Caenorhabditis elegans, the distal tip cell/germline interaction promotes a mitotic fate and/or inhibits meiosis through GLP-1/Notch signaling. However, GLP-1-mediated signaling alone is not sufficient for a wild-type level of germline proliferation. Here, we provide evidence that specific cells of the somatic gonadal sheath lineage influence amplification, differentiation, and the potential for tumorigenesis of the germ line. First, an interaction between the distal-most pair of sheath cells and the proliferation zone of the germ line is required for larval germline amplification. Second, we show that insufficient larval germline amplification retards gonad elongation and thus delays meiotic entry. Third, a more severe delay in meiotic entry, as is exhibited in certain mutant backgrounds, inappropriately juxtaposes undifferentiated germ cells with cells of the proximal sheath lineage, leading to the formation of a proximal germline tumor derived from undifferentiated germ cells. Tumors derived from dedifferentiated germ cells, however, respond to the proximal interaction differently depending on the mutant background. Our study underscores the importance of strict developmental coordination between neighboring tissues. We discuss these results in the context of mechanisms that may underlie tumorigenesis.

5. Maciejowski, John, Ahn, James Hyungsoo, Cipriani, Patricia Giselle, Killian, Darrell J, Chaudhary, Aisha L, Lee, Ji Inn, Voutev, Roumen, Johnsen, Robert C, Baillie, David L, Gunsalus, Kristin C, Fitch, David H A, Hubbard, E Jane Albert. "Autosomal genes of autosomal/X-linked duplicated gene pairs and germ-line proliferation in Caenorhabditis elegans," Genetics 2005 Apr;169(4):1997-2011. Epub 2005 Jan 31.   (MEDL:15687263 PMID: 15687263 #J0126777)    

   We report molecular genetic studies of three genes involved in early germ-line proliferation in Caenorhabditis elegans that lend unexpected insight into a germ-line/soma functional separation of autosomal/X-linked duplicated gene pairs. In a genetic screen for germ-line proliferation-defective mutants, we identified mutations in rpl-11.1 (L11 protein of the large ribosomal subunit), pab-1 [a poly(A)-binding protein], and glp-3/eft-3 (an elongation factor 1-alpha homolog). All three are members of autosome/X gene pairs. Consistent with a germ-line-restricted function of rpl-11.1 and pab-1, mutations in these genes extend life span and cause gigantism. We further examined the RNAi phenotypes of the three sets of rpl genes (rpl-11, rpl-24, and rpl-25) and found that for the two rpl genes with autosomal/X-linked pairs (rpl-11 and rpl-25), zygotic germ-line function is carried by the autosomal copy. Available RNAi results for highly conserved autosomal/X-linked gene pairs suggest that other duplicated genes may follow a similar trend. The three rpl and the pab-1/2 duplications predate the divergence between C. elegans and C. briggsae, while the eft-3/4 duplication appears to have occurred in the lineage to C. elegans after it diverged from C. briggsae. The duplicated C. briggsae orthologs of the three C. elegans autosomal/X-linked gene pairs also display functional differences between paralogs. We present hypotheses for evolutionary mechanisms that may underlie germ-line/soma subfunctionalization of duplicated genes, taking into account the role of X chromosome silencing in the germ line and analogous mammalian phenomena.

6. Hansen, Dave, Hubbard, E Jane Albert, Schedl, Tim. "Multi-pathway control of the proliferation versus meiotic development decision in the Caenorhabditis elegans germline," Developmental biology (Orlando) 2004 Apr 15;268(2):342-57.   (MEDL:15063172 PMID: 15063172 #J0126779)    

   An important event in the development of the germline is the initiation of meiotic development. In Caenorhabditis elegans, the conserved GLP-1/Notch signaling pathway regulates the proliferative versus meiotic entry decision, at least in part, by spatially inhibiting genes in the gld-1 and gld-2 parallel pathways, which are proposed to either inhibit proliferation and/or promote meiotic development. Mutations that cause constitutive activation of the GLP-1 pathway, or inactivation of both the gld-1 and gld-2 parallel pathways, result in a tumorous germline in which all cells are thought to be proliferative. Here, to analyze proliferation and meiotic entry in wild-type and mutant tumorous germlines, we use anti-REC-8 and anti-HIM-3 specific antibodies as markers, which under our fixation conditions, stain proliferative and meiotic cells, respectively. Using these makers in wild-type animals, we find that the border of the switch from proliferation to meiotic entry is staggered in late-larval and adult germlines. In wild-type adults, the switch occurs between 19 and 26 cell diameters from the distal end, on average. Our analysis of mutants reveals that tumorous germlines that form when GLP-1 is constitutively active are completely proliferative, while tumors due to inactivation of the gld-1 and gld-2 pathways show evidence of meiotic entry. Genetic and time course studies suggest that a third pathway may exist, parallel to the GLD-1 and GLD-2 pathways, that promotes meiotic development.

7. Killian, Darrell J, Hubbard, E Jane Albert. "C. elegans pro-1 activity is required for soma/germline interactions that influence proliferation and differentiation in the germ line," Development 2004 Mar;131(6):1267-78. Epub 2004 Feb 18.   (MEDL:14973273 PMID: 14973273 #J0126780)    

   Strict spatial and temporal regulation of proliferation and differentiation is essential for proper germline development and often involves soma/germline interactions. In C. elegans, a particularly striking outcome of defective regulation of the proliferation/differentiation pattern is the Pro phenotype in which an ectopic mass of proliferating germ cells occupies the proximal adult germ line, a region normally occupied by gametes. We describe a reduction-of-function mutation in the gene pro-1 that causes a highly penetrant Pro phenotype. The pro-1 mutant Pro phenotype stems from defects in the time and position of the first meiotic entry during early germline development. pro-1(RNAi) produces a loss of somatic gonad structures and concomitant reduction in germline proliferation and gametogenesis. pro-1 encodes a member of a highly conserved subfamily of WD-repeat proteins. pro-1(+) is required in the sheath/spermatheca lineage of the somatic gonad in its role in the proper establishment of the proliferation/differentiation pattern in the germline. Our results provide a handle for further analysis of this soma-to-germline interaction.

8. Hubbard, E Jane Albert, Pera, Renee A Reijo. "A germ-cell odyssey: fate, survival, migration, stem cells and differentiation. Meeting on germ cells," EMBO reports 2003 Apr;4(4):352-7. Epub 2003 Mar 21.   (MEDL:12671678 PMID: 12671678 #J0126782)    

9. Mishra, Bud, Daruwala, Raoul-Sam, Zhou, Yi, Ugel, Nadia, Policriti, Alberto, Antoniotti, Marco, Paxia, Salvatore, Rejali, Marc, Rudra, Archisman, Cherepinsky, Vera, Silver, Naomi, Casey, William, Piazza, Carla, Simeoni, Marta, Barbano, Paolo, Spivak, Marina, Feng, Jiawu, Gill, Ofer, Venkatesh, Mysore, Cheng, Fang, Sun, Bing, Ioniata, Iuliana, Anantharaman, Thomas, Hubbard, E Jane Albert, Pnueli, Amir, Harel, David, Chandru, Vijay, Hariharan, Ramesh, Wigler, Michael, Park, Frank, Lin, Shih-Chieh, Lazebnik, Yuri, Winkler, Franz, Cantor, Charles R, Carbone, Alessandra, Gromov, Mikhael. "A sense of life: computational and experimental investigations with models of biochemical and evolutionary processes," Omics : a journal of integrative biology 2003 Fall;7(3):253-68.   (MEDL:14583115 PMID: 14583115 #J0125778)    

   We collaborate in a research program aimed at creating a rigorous framework, experimental infrastructure, and computational environment for understanding, experimenting with, manipulating, and modifying a diverse set of fundamental biological processes at multiple scales and spatio-temporal modes. The novelty of our research is based on an approach that (i) requires coevolution of experimental science and theoretical techniques and (ii) exploits a certain universality in biology guided by a parsimonious model of evolutionary mechanisms operating at the genomic level and manifesting at the proteomic, transcriptomic, phylogenic, and other higher levels. Our current program in 'systems biology' endeavors to marry large-scale biological experiments with the tools to ponder and reason about large, complex, and subtle natural systems. To achieve this ambitious goal, ideas and concepts are combined from many different fields: biological experimentation, applied mathematical modeling, computational reasoning schemes, and large-scale numerical and symbolic simulations. From a biological viewpoint, the basic issues are many: (i) understanding common and shared structural motifs among biological processes; (ii) modeling biological noise due to interactions among a small number of key molecules or loss of synchrony; (iii) explaining the robustness of these systems in spite of such noise; and (iv) cataloging multistatic behavior and adaptation exhibited by many biological processes.

10. Pepper, Anita S-R, Lo, Te Wen, Killian, Darrell J, Hall, David H, Hubbard, E Jane Albert. "The establishment of Caenorhabditis elegans germline pattern is controlled by overlapping proximal and distal somatic gonad signals," Developmental biology (Orlando) 2003 Jul 15;259(2):336-50.   (MEDL:12871705 PMID: 12871705 #J0126781)    

    We investigated the control of proliferation and differentiation in the larval Caenorhabditis elegans hermaphrodite germ line through analysis of glp-1 and lag-2 mutants, cell ablations, and ultrastructural data. After the first several rounds of germ cell division, GLP-1, a receptor of the LIN-12/Notch family, governs germline proliferation. We analyzed the proximal proliferation (Pro) phenotype in glp-1(ar202) and found that initial meiosis was delayed and spatially mispositioned. This is due, at least in part, to a heightened response of the mutant GLP-1 receptor to multiple sources of the somatic ligand LAG-2, including the proximal somatic gonad. We investigated whether proximal LAG-2 affects germline proliferation in the wild type. Our results indicate that (1) LAG-2 is necessary for GLP-1-mediated germline proliferation and prevention of early meiosis, and (2) several distinct anatomical sources of LAG-2 in the larval somatic gonad functionally overlap to promote proliferation and prevent early meiosis. Ultrastructural studies suggest that mitosis is not restricted to areas of direct DTC-germ line contact and that the germ line shares a common cytoplasm in larval stages. We propose that downregulation of the GLP-1 signaling pathway in the proximal germ line at the time of meiotic onset is under tight temporal and spatial control.

11. Pepper, Anita S-R, Killian, Darrell J, Hubbard, E Jane Albert. "Genetic analysis of Caenorhabditis elegans glp-1 mutants suggests receptor interaction or competition," Genetics 2003 Jan;163(1):115-32.   (MEDL:12586701 PMID: 12586701 #J0126783)    

    glp-1 encodes a member of the highly conserved LIN-12/Notch family of receptors that mediates the mitosis/meiosis decision in the C. elegans germline. We have characterized three mutations that represent a new genetic and phenotypic class of glp-1 mutants, glp-1(Pro). The glp-1(Pro) mutants display gain-of-function germline pattern defects, most notably a proximal proliferation (Pro) phenotype. Each of three glp-1(Pro) alleles encodes a single amino acid change in the extracellular part of the receptor: two in the LIN-12/Notch repeats (LNRs) and one between the LNRs and the transmembrane domain. Unlike other previously described gain-of-function mutations that affect this region of LIN-12/Notch family receptors, the genetic behavior of glp-1(Pro) alleles is not consistent with simple hypermorphic activity. Instead, the mutant phenotype is suppressed by wild-type doses of glp-1. Moreover, a trans-heterozygous combination of two highly penetrant glp-1(Pro) mutations is mutually suppressing. These results lend support to a model for a higher-order receptor complex and/or competition among receptor proteins for limiting factors that are required for proper regulation of receptor activity. Double-mutant analysis with suppressors and enhancers of lin-12 and glp-1 further suggests that the functional defect in glp-1(Pro) mutants occurs prior to or at the level of ligand interaction.

12. Hubbard, E J, Greenstein, D. "The Caenorhabditis elegans gonad: a test tube for cell and developmental biology," Developmental dynamics 2000 May;218(1):2-22.   (MEDL:10822256 PMID: 10822256 #J0126813)    

    Sexual reproduction of multicellular organisms depends critically on the coordinate development of the germ line and somatic gonad, a process known as gonadogenesis. Together these tissues ensure the formation of functional gametes and, in the female of many species, create a context for production and further development of the zygote. Since the future of the species hangs in the balance, it is not surprising that gonadogenesis is a complex process involving conserved and multi-faceted developmental mechanisms. Genetic, anatomical, cell biological, and molecular experiments have established the nematode Caenorhabditis elegans as a paradigm for studying gonadogenesis. Furthermore, these studies demonstrate the utility of C. elegans gonadogenesis for exploring broad issues in cell and developmental biology, such as cell fate specification, morphogenesis, cell signaling, cell cycle control, and programmed cell death. The synergy of molecular genetics and cell biology conducted at single-cell resolution in real time permits an extraordinary depth of analysis in this organism. In this review, we first describe the embryonic and post-embryonic development and morphology of the C. elegans gonad. Next we recount seminal experiments that established the field, highlight recent results that provide insight into conserved developmental mechanisms, and present future prospects for the field.

13. Hubbard, E J, Wu, G, Kitajewski, J, Greenwald, I. "sel-10, a negative regulator of lin-12 activity in Caenorhabditis elegans, encodes a member of the CDC4 family of proteins," Genes & development 1997 Dec 1;11(23):3182-93.   (MEDL:9389650 PMID: 9389650 #J0126814)    

    Mutations that influence lin-12 activity in Caenorhabditis elegans may identify conserved factors that regulate the activity of lin-12/Notch proteins. We describe genetic evidence indicating that sel-10 is a negative regulator of lin-12/Notch-mediated signaling in C. elegans. Sequence analysis shows that SEL-10 is a member of the CDC4 family of proteins and has a potential human ortholog. Coimmunoprecipitation data indicate that C. elegans SEL-10 complexes with LIN-12 and with murine Notch4. We propose that SEL-10 promotes the ubiquitin-mediated turnover of LIN-12/Notch proteins, and discuss potential roles for the regulation of lin-12/Notch activity by sel-10 in cell fate decisions and tumorigenesis.

14. Hubbard, E J, Dong, Q, Greenwald, I. "Evidence for physical and functional association between EMB-5 and LIN-12 in Caenorhabditis elegans," Science 1996 Jul 5;273(5271):112-5.   (MEDL:8658178 PMID: 8658178 #J0126815)    

    The Caenorhabditis elegans LIN-12 and GLP-1 proteins are members of the LIN-12/Notch family of receptors for intercellular signals that specify cell fate. Evidence presented here suggests that the intracellular domains of LIN-12 and GLP-1 interact with the C. elegans EMB-5 protein and that the emb-5 gene functions in the same pathway as the lin-12 and glp-1 genes. EMB-5 is similar in sequence to a yeast protein that controls chromatin structure. Hence, a direct consequence of LIN-12 or GLP-1 activation may be an alteration of chromatin structure that produces changes in transcriptional activity.

15. Hubbard, E J, Yang, X L, Carlson, M. "Relationship of the cAMP-dependent protein kinase pathway to the SNF1 protein kinase and invertase expression in Saccharomyces cerevisiae," Genetics 1992 Jan;130(1):71-80.   (MEDL:1310088 PMID: 1310088 #J0126818)    

    The SNF1 protein kinase and the associated SNF4 protein are required for release of glucose repression in Saccharomyces cerevisiae. To identify functionally related proteins, we selected genes that in multicopy suppress the raffinose growth defect of snf4 mutants. Among the nine genes recovered were two genes from the cAMP-dependent protein kinase (cAPK) pathway, MSI1 and PDE2. Increased dosage of these genes partially compensates for defects in nutrient utilization and sporulation in snf1 and snf4 null mutants, but does not restore invertase expression. These results suggest that SNF1 and cAPK affect some of the same cellular responses to nutrients. To examine the role of the cAPK pathway in regulation of invertase, we assayed mutants in which the cAPK is not modulated by cAMP. Expression of invertase was regulated in response to glucose and was dependent on SNF1 function. Thus, a cAMP-responsive cAPK is dispensable for regulation of invertase.

16. Robinson, L C, Hubbard, E J, Graves, P R, DePaoli-Roach, A A, Roach, P J, Kung, C, Haas, D W, Hagedorn, C H, Goebl, M, Culbertson, M R. "Yeast casein kinase I homologues: an essential gene pair," Proceedings of the National Academy of Sciences of the United States of America 1992 Jan 1;89(1):28-32.   (MEDL:1729698 PMID: 1729698 #J0126817)    

    We report the isolation of an essential pair of Saccharomyces cerevisiae genes that encode protein kinase homologues. The two genes were independently isolated as dosage-dependent suppressors. Increased dosage of YCK1 suppressed defects caused by reduced SNF1 protein kinase activity, and increased dosage of YCK2 relieved sensitivity of wild-type cells to salt stress. The two genes function identically in the two growth assays, and loss of function of either gene alone has no discernible effect on growth. However, loss of function of both genes results in inviability. The two predicted protein products share 77% overall amino acid identity and contain sequence elements conserved among protein kinases. Partial sequence obtained for rabbit casein kinase I shares 64% identity with the two yeast gene products. Moreover, an increase in casein kinase I activity is observed in extracts from cells overexpressing YCK2. Thus YCK1 and YCK2 appear to encode casein kinase I homologues.

17. Yang, X, Hubbard, E J, Carlson, M. "A protein kinase substrate identified by the two-hybrid system," Science 1992 Jul 31;257(5070):680-2.   (MEDL:1496382 PMID: 1496382 #J0126816)    

    A genetic method, the two-hybrid system, was used to identify four genes encoding proteins that interact with the SNF1 protein kinase from yeast. One of the genes, SIP1, was independently isolated as a multicopy suppressor of defects caused by reduced SNF1 kinase activity, and genetic evidence supports its function in the SNF1 pathway. The SIP1 protein co-immunoprecipitated with SNF1 and was phosphorylated in vitro. Thus, the two-hybrid system, which is applicable to any cloned gene, can be used to detect physical interactions between protein kinases and functionally related substrate proteins.