Date Published: February 16, 2017
Publisher: Public Library of Science
Author(s): Zachary T. Neeb, Daniel J. Hogan, Sol Katzman, Alan M. Zahler, Emanuele Buratti.
During its sexual reproduction, the stichotrichous ciliate Oxytricha trifallax orchestrates a remarkable transformation of one of the newly formed germline micronuclear genomes. Hundreds of thousands of gene pieces are stitched together, excised from chromosomes, and replicated dozens of times to yield a functional somatic macronuclear genome composed of ~16,000 distinct DNA molecules that typically encode a single gene. Little is known about the proteins that carry out this process. We profiled mRNA expression as a function of macronuclear development and identified hundreds of mRNAs preferentially expressed at specific times during the program. We find that a disproportionate number of these mRNAs encode proteins that are involved in DNA and RNA functions. Many mRNAs preferentially expressed during macronuclear development have paralogs that are either expressed constitutively or are expressed at different times during macronuclear development, including many components of the RNA polymerase II machinery and homologous recombination complexes. Hundreds of macronuclear development-specific genes encode proteins that are well-conserved among multicellular eukaryotes, including many with links to germline functions or development. Our work implicates dozens of DNA and RNA-binding proteins with diverse evolutionary trajectories in macronuclear development in O. trifallax. It suggests functional connections between the process of macronuclear development in unicellular ciliates and germline specialization and differentiation in multicellular organisms, and argues that gene duplication is a key source of evolutionary innovation in this process.
Ciliates are diverse, abundant and extremely successful unicellular eukaryotes that display a special case of germline-soma specialization vis-à-vis nuclear dimorphism; a germline nucleus (micronucleus) used for propagation of genetic information and a somatic nucleus (macronucleus) used for cell growth . When starved, cells of different mating types pair, micronuclei undergo meiosis, they exchange haploid micronuclei which fuse to form a new diploid micronucleus, they perform one to several rounds of micronuclear mitosis, and then develop a new macronucleus from one of the newly formed micronuclei. This differentiation program is associated with reorganization of the genome and in some cases extraordinary genome rearrangements. In all ciliate lineages studied, genome rearrangements are epigenetically determined by communication of DNA content between macronuclei and micronuclei via RNA intermediates [2–4]. Several factors intrinsic to germline specialization and differentiation were first characterized in ciliates, such as telomerase  and histone acetyl transferase , as well as seminal studies on specialized histones, PIWI and HP1 proteins [7–19]. Thus, ciliates provide relatively simple and facile systems to study principles of germline-soma specialization, germline differentiation and RNA-mediated epigenetic memory.
Identifying the mechanisms by which the single-celled ciliate O. trifallax evolved two remarkable genomes segregated for germline and somatic functions and transforms the scrambled, nonfunctional germline genome into the highly streamlined functional somatic genome after sexual reproduction would provide insights into the principles underlying evolutionary innovation. The mRNA expression profiles of macronuclear development in O. trifallax presented herein implicate a staggering array of DNA and RNA-interacting proteins in this process. From our work, we hypothesize that the specialized function of genes involved in macronuclear development is acquired via several mechanisms: (i) a core set of factors involved in both macronuclear development across ciliates and germline-soma stratification in multicellular organisms (Fig 4), (ii) expansion of protein families with domains involved in DNA and RNA metabolism (Fig 1), (iii) specialization of multimember DNA and RNA-binding protein machines via gene duplication (Figs 2 and 3) and (iv) recent acquisition of diverse DNA and RNA remodeling factors, such as through transposon domestication (Fig 5) [26, 85].