Mary Lou King, Ph.D.
Germline Determination in the Xenopus Embryo
The exclusion of germ cells from somatic cell fates in early development is an essential process in metazoans that ensures continuation of the species. Primordial germ cells (PGCs), precursors to the gametes, are specified through the inheritance of a sub-cellular domain called the germ plasm. Components of the germ plasm are responsible for at least four activities that are required to both protect PGCs from somatic differentiation and to initiate the unique gene expression programs of the germline: 1) activation of sequestered maternal germline mRNAs; 2) repression of maternal somatic messages; 3) transient genome-wide suppression of transcription to ensure that somatic programs are not activated when zygotic transcription initiates in the rest of the embryo; and 4) transcriptional activation of PGC-specific genes after degradation of maternal somatic mRNAs. All of these activities occur in the absence of transcription and therefore must be regulated primarily at the level of translation.
The goal of my research program is to define key players in this post-transcriptional regulatory network that protects and specifies the germline. The RNA-binding proteins Nanos and Dazl are translational regulators in the germlines of diverse species, including frogs and humans. Both are translationally activated in PGCs by an unknown mechanism. Nanos and Dazl both interact with another (sequence-specific) RNA-binding protein, Pumilio (Pum), to regulate translation, but with opposite effects: Nanos represses translation of target RNAs, while Dazl promotes it. Our work in Xenopus has shown that PGCs lacking Nanos prematurely initiate Pol II transcription, inappropriately express somatic genes, and do not survive. Xenopus PGCs deficient in dazl fail to migrate to the primordial gonads and are lost from the germline. We are currently addressing the key questions: What activates nanos and dazl translation? Preliminary studies support a new role for the RNA-binding protein, Dead-end, in this process. What are the identities of the target mRNAs that Nanos, Xdazl, and Dead-end regulate?
Identification of proteins and RNAs in the germ plasm
Xenopus offers a unique opportunity to isolate biochemical quantities of germ plasm, making it possible, with current technology, to create a complete “parts list” of this cellular “machine” that specifies the totipotent germ cell lineage. Moreover, Xenopus is highly amenable to expression cloning, allowing a high-throughput approach to assess gene function of germ plasm components. In a parallel set of studies, we have enriched for germ plasm from two different stages in quantities sufficient for protein analyses by Liquid Chromatography-tandem Mass Spectrometry and for RNA identification by RNA sequencing. RNA-seq analyses are being done for coding, long non-coding and micro-RNAs. The results from these studies should reveal gene networks that regulate totipotency and immortality, two hallmarks of the germline.
Owens DA, Butler AM, Aguero TH, Newman KM, Van Booven D, King ML. (2017). “High-throughput analysis reveals novel maternal germline RNAs crucial for primordial germ cell preservation and proper migration”. Development. 144(2):292-304. doi: 10.1242/dev.139220.
King ML. (2017) “Maternal messages to live by: a personal historical perspective”. Genesis. 55(1-2). doi: 10.1002/dvg.23007. Review.
Aguero T, Kassmer S, Alberio R, Johnson A, King ML. (2017) “Mechanisms of Vertebrate Germ Cell Determination”. Adv Exp Med Biol. 953:383-440
Butler AM, Aguero T, Newman KM, King ML. (2017) “Primordial Germ Cell Isolation from Xenopus laevis Embryos”. Methods Mol Biol.1463:115-124.
Aguero T, Zhou Y, Kloc M, Chang P, Houliston E, King ML. (2016) “Hermes (Rbpms) is a Critical Component of RNP Complexes that Sequester Germline RNAs during Oogenesis”. J Dev Biol.4(1). pii: 2.
Yang, J, Aguero, T., and King, M.L. (2015). “The Xenopus Maternal to Zygotic Transition from the Perspective of the Germline “. In: Current Topics in Developmental Biology: “The Maternal-to-Zygotic Transition” (ed. Howard Lipshitz). Academic Press, N.Y. Accepted.
Aguero T., Kassmer, S., Alberio, R., Johnson, A., and King, M.L. (2015). “Mechanisms of Vertebrate Germ Cell Determination”, Ch 8. In “Early Vertebrate Development: The Egg-to-Embryo Transition”, (eds. Francisco Pelegri, Michael Danilchik and Ann E. Sutherland). Springer Press. Accepted.
King M.L. (2014) “Germ cell Specification in Xenopus” in “Xenopus Development” pp. 75-100. ed by M. Kloc and J.Z. Kubiak. John Wiley & Sons, Inc. Hoboken, New Jersey.
Lai, Fangfang and King, M.L. (2013) Repressive translational control in germ cells. Mol. Repro. Dev.,80(8):665-76. Review.
King M.L. (2013) Invited review “Germ cell Specification” in “Xenopus Development” ed by M. Kloc and J.Z. Kubiak. John Wiley & Sons, Inc. In press.
Wenyan Mei, Zhigang Jin, Fangfang Lai, Tyler Schwend, Douglas W. Houston, Mary Lou King, Jing Yang (2013). Maternal Dead-End1 is required for vegetal cortical microtubule assembly during Xenopus axis specification. Development,140, 2334-44.
Fangfang Lai, Amar Singh, and Mary Lou King. (2012) Xenopus Nanos1 is required to Prevent Endoderm Gene Expression, Transcription and Apoptosis in Primordial Germ Cells. Development, 139, 1476-1486.
Hua-wei Wang, Jun-shun Fang, Xia Kuang, Li-yun Miao,Guo-liang Xia, Mary Lou King, Jian Zhang (2012). Long-chain acyl-CoA synthetase 1 is required for maintaining meiotic arrest in Xenopus and mouse oocytes. Biology of Reproduction, 87,1-9.
Luo, X., Nerlick, S., An, W., King, M.L. (2011). Xenopus Germline nanos1 is Translationally Repressed by a Novel Structure Bsed Mechanism. Development, 138:589-98.
Lai, F, Zhou, Y., Luo, X., Fox, J., and *King, M.L. * (2011). Nanos1 functions as a Translational Repressor in the Xenopus Germline. Mech Dev 128:153-63.
Venkatarama, T., Lai, F., Luo, X., Zhou, Y., Newman, K., King, M.L. (2010). Repression of Zygotic Gene Expression in the Xenopus Germline. Development, 137:651-660.
Rodrigues, C.O., Nerlick, S., White, E.L., Cleveland, J.L. and King, M.L. (2008). A Myc-Slug (Snail2)/Twist Regulatory Circuit Directs Vascular Development. Development, 135:1903-11.
Song, H-W., Cauffman, K., Chan, A.P., Zhou, Y., *King, M.L., *Etkin, L.D. and Kloc, M. (2007). Hermes RNA binding protein targets RNAs encoding proteins involved in meiotic maturation, early cleavage, and germline development. Differentiation, 75:519-28.
*King, M. L., *Messitt, and Mowry, K. (2005) Putting RNAs in the Right Place at the Right Time: RNA Localization in the Frog Oocyte. Biol. Cell 97: 19-33.
Machado RJ, Moore W, Hames R, Houliston E, Chang P, King ML, Woodland HR. (2005) Xenopus Xpat protein is a major component of germ plasm and may function in its organization and positioning. Dev Biol. 287(2):289-300.
Chang, P., Torres, J., Lewis, R., Mowry, K., Houlison, E and King, M. L. (2004) Localization of RNAs to the mitochondrial cloud in Xenopus oocytes by entrapment and association with endoplasmic reticulum. Mol. Biol. Cell 15:4669-4681.
Zhou, Y., Zhang, J. and King, M.L. (2003) Xenopus ARH Couples Lipoprotein Receptors with the AP-2 Complex in Oocytes and Embryos and is Required for Vitellogenesis. J Biological Chemistry 278:44584-44592.
Bubunenko, M., Kress, T.L., Vempati, U.D., Mowry, K. King, M.L. (2002) A consensus RNA signal that directs germ layer determinants to the vegetal cortex of Xenopus oocytes. Dev. Biol. 248:82-92.
Houston, D. W. and King, M.L. (2000) A critical role for Xdazl, a germ plasm-localized RNA, in the differentiation of primordial germ cells in Xenopus. Development 127:447-456.
View published research articles by Dr. King in the National Library of Medicine.