The nature and origin of nucleus-like intracellular inclusions in Paleoproterozoic eukaryote microfossils K. PANG, 1,2 Q. TANG, 1,2 J. D. SCHIFFBAUER, 3 J. YAO, 4 X. YUAN, 1 B. WAN, 1 L. CHEN, 1 Z. OU 1 AND S. XIAO 2 1 State Key Laboratory of Paleobiology and Stratigraphy, Nanjing Institute of Geology and Paleontology, Chinese Academy of Sciences, Nanjing, China 2 Department of Geosciences, Virginia Polytechnic Institute and State University, Blacksburg, VA, USA 3 Department of Geological Sciences, University of Missouri, Columbia, MO, USA 4 College of Life Sciences, Peking University, Beijing, China ABSTRACT The well-known debate on the nature and origin of intracellular inclusions (ICIs) in silicified microfossils from the early Neoproterozoic Bitter Springs Formation has recently been revived by reports of possible fos- silized nuclei in phosphatized animal embryo-like fossils from the Ediacaran Doushantuo Formation of South China. The revisitation of this discussion prompted a critical and comprehensive investigation of ICIs in some of the oldest indisputable eukaryote microfossils—the ornamented acritarchs Dictyosphaera delicata and Shuiyousphaeridium macroreticulatum from the Paleoproterozoic Ruyang Group of North China— using a suite of characterization approaches: scanning electron microscopy (SEM), transmission electron microscopy (TEM), and focused ion beam scanning electron microscopy (FIB-SEM). Although the Ruyang acritarchs must have had nuclei when alive, our data suggest that their ICIs represent neither fossilized nuclei nor taphonomically condensed cytoplasm. We instead propose that these ICIs likely represent biolog- ically contracted and consolidated eukaryotic protoplasts (the combination of the nucleus, surrounding cytoplasm, and plasma membrane). As opposed to degradational contraction of prokaryotic cells within a mucoidal sheath—a model proposed to explain the Bitter Springs ICIs—our model implies that protoplast condensation in the Ruyang acritarchs was an in vivo biologically programmed response to adverse condi- tions in preparation for encystment. While the discovery of bona fide nuclei in Paleoproterozoic acritarchs would be a substantial landmark in our understanding of eukaryote evolution, the various processes (such as degradational and biological condensation of protoplasts) capable of producing nuclei-mimicking struc- tures require that interpretation of ICIs as fossilized nuclei be based on comprehensive investigations. Received 4 May 2013; accepted 23 July 2013 Corresponding author: S. Xiao. Tel.: +1 540 231 1366; fax: +1 540 231 3386; e-mail: xiao@vt.edu INTRODUCTION Recognized as one of the major evolutionary transitions in the history of life (Maynard Smith & Szathm ary, 1997), the rise of eukaryotes is an important milestone in evolu- tion and paves the road for subsequent milestones, includ- ing the origin of complex multicellularity (Knoll, 2011). Reliably interpreting the fossil record of early eukaryotes, however, is highly complicated. Steranes, which are regarded as derivatives from eukaryote-specific sterols, have been reported from ~2700 Ma shales in northwestern Australia (Brocks et al., 1999), but their origin and syngenicity remain unresolved (Sherman et al., 2007; Ras- mussen et al., 2008). Cell size was once regarded as a use- ful criterion for distinguishing fossil eukaryotes (Schopf & Oehler, 1976), but the considerable overlap between eukaryote and bacterial cell size ranges means that this cri- terion alone is only suggestive, not conclusive, evidence for a eukaryotic affinity. Recently, cell wall ultrastructures have been used in combination with vesicle ornamentation and extravesicular processes as evidence for eukaryotic organisms (Javaux et al., 2003, 2004; Knoll et al., 2006; Moczydlowska & Willman, 2009). However, because ultra- structures of extant cellular life have not been exhaustively © 2013 John Wiley & Sons Ltd 499 Geobiology (2013), 11, 499–510 DOI: 10.1111/gbi.12053