Visualization of Chromatin in the Yeast Nucleus and Nucleolus Using Hyperosmotic Shock.

Nicolas Thelen,Marc Thiry,Denis L. J. Lafontaine,Jean Defourny

doi:10.3390/ijms22031132

Abstract

Unlike in most eukaryotic cells, the genetic information of budding yeast in the exponential growth phase is only present in the form of decondensed chromatin, a configuration that does not allow its visualization in cell nuclei conventionally prepared for transmission electron microscopy. In this work, we studied the distribution of chromatin and its relationships to the nucleolus using different cytochemical and immunocytological approaches applied to yeast cells subjected to hyperosmotic shock. Our results show that osmotic shock induces the formation of heterochromatin patches in the nucleoplasm and intranucleolar regions of the yeast nucleus. In the nucleolus, we further revealed the presence of osmotic shock-resistant DNA in the fibrillar cords which, in places, take on a pinnate appearance reminiscent of ribosomal genes in active transcription as observed after molecular spreading (“Christmas trees”). We also identified chromatin-associated granules whose size, composition and behaviour after osmotic shock are reminiscent of that of mammalian perichromatin granules. Altogether, these data reveal that it is possible to visualize heterochromatin in yeast and suggest that the yeast nucleus displays a less-effective compartmentalized organization than that of mammals.

Highlights

The eukaryotic cell is characterised by the presence of a compartment bounded by a double membrane, called the nucleus, which contains genetic information
We studied the distribution of chromatin in the yeast nucleus at the ultrastructural level and its relationships to the nucleolus using different cytochemical and immunocytological techniques
In exponentially growing yeast cells conventionally prepared for transmission electron microscopy, we showed that the vast majority of nuclear chromatin does not form a condensed structure

Summary

Introduction

The eukaryotic cell is characterised by the presence of a compartment bounded by a double membrane, called the nucleus, which contains genetic information The latter is organised into chromatin domains with distinct structure and function [1,2]. Heterochromatin is usually condensed and lacks highly expressed genes [3] It concentrates mainly at the periphery of the nucleus and around the nucleolus. The nucleus contains various membraneless subcompartments formed by liquid–liquid phase separation (LLPS), which concentrates specific proteins and excludes others to form RNA and protein-rich microenvironments that promote or inhibit certain activities. These are referred to as biomolecular condensates. Recent data support the notion that the nucleolus is a multilayered biomolecular condensate whose formation by LLPS facilitates the initiation of the first stages of ribosome biogenesis and other functions [6]

Methods

Results

Conclusion