Dynein-Based Accumulation of Membranes Regulates Nuclear Expansion in Xenopus laevis Egg Extracts

Abstract

Nuclear size changes dynamically during development and has long been observed to correlate with the space surrounding the nucleus, as well as with the volume of the cell. Here we combine an in vitro cell-free system of Xenopus laevis egg extract with microfluidic devices to systematically analyze the effect of spatial constraints. The speed of nuclear expansion depended on the available space surrounding the nucleus up to a threshold volume in the nanoliter range, herein referred to as the nuclear domain. Under spatial constraints smaller than this nuclear domain, the size of microtubule-occupied space surrounding the nucleus turned out to be limiting for the accumulation of membranes around the nucleus via the motor protein dynein, therefore determining the speed of nuclear expansion. This mechanism explains how spatial information surrounding the nucleus, such as the positioning of the nucleus inside the cell, can control nuclear expansion.

Turn Up the Volume: Uncovering Nucleus Size Control Mechanisms

“…Similar scaling relationships have been reported for the mitotic spindle (Good et al, 2013) and centrosome (Decker et al, 2011), suggesting that cytoplasmic volume may be a universal regulator of organelle growth (Goehring and Hyman, 2012). Hara and Merten (2015) also identified molecular factors, including the amount of membrane and the levels of the motor protein dynein, that contribute to the rate of nucleus expansion. Inhibition of microtubule assembly or dynein activity restricted the growth of nuclei both in test tubes and while encapsulated in microfabricated channels.…”

“…For example, do changes in nucleus size directly alter transcription or the spatial organization of the genome? Given that an altered nucleocytoplasmic ratio is linked to cellular senescence and cancer, in vitro studies like this one by Hara and Merten (2015) may provide paradigms for understanding size control relevant to both healthy and diseased cells. Growth is dependent on the amount of locally accessible cytoplasm.…”

“…In a study published in this issue of Developmental Cell, Hara and Merten (2015) develop and apply an in vitro reconstitution approach-combining cellfree cytoplasmic extracts and confinement-to uncover relationships between the cytoplasmic space surrounding the nucleus and nuclear growth ( Figure 1A). The power of their experimental system is that individual parameters, such as compartment size and membrane concentration, can be studied in isolation and without adverse effects on the cell or embryo.…”

Nuclei and nucleoli in embryo-like fossils from the Ediacaran Weng’an Biota

“…This interpretation was subsequently rejected on the grounds that the LISs are unfeasibly large, and the cells of later developmental stages are smaller, too small to contain the same amount of genetic material . However, given cell size, the nuclei are not unfeasibly large, or even unusually large for palintomically dividing cells (Huldtgren et al, 2012), and analyses have demonstrated an isometric relationship between cell and nucleus volume in living eukaryotes, like the one that we observe (Cavalier-Smith, 2005;Conklin, 1912;Goehring and Hyman, 2012;Kimura, 2009, 2011;Hara and Merten, 2015;Jorgensen et al, 2007;Neumann and Nurse, 2007;Tsichlaki and FitzHarris, 2016;Wilson, 1925). Furthermore, in living eukaryote systems, within a few rounds of palintomy, cell volume does indeed diminish to less than the original volume of the nucleus (Tsichlaki and FitzHarris, 2016).…”

Recent advances in understanding nuclear size and shape

“…Additionally, when X. laevis egg extract was treated with a dynein inhibitor prior to encapsulation in the microchannels, nuclear expansion was greatly inhibited, implicating a microtubuleand dynein-based mechanism of nuclear size regulation. 137 Other new technologies allow for measuring the entire transcriptomes of individual cells, by coupling microfluidic encapsulation of individual cells with next generation sequencing. 138 For example, whereas only 5 mouse retinal cell types were previously known, this so-called Drop-seq method identified 39 distinct cell types based on individual cell profiles.…”