Jezek 2011 AbstractMitoComLectures

Jezek P, Dlaskova A, Santorova J, Spacek T, Smolkova K, Tauber J, Mlodzianoski MJ, Bewersdorf J (2011) 3D visualization of mitochondrial network and mtDNA nucleoids at 30 nm by biplane FPALM microscopy. Abstract MitoCom Lectures.

» [Lectures]

Jezek P, Dlaskova A, Santorova J, Spacek T, Smolkova K, Tauber J, Mlodzianoski MJ, Bewersdorf J (2011) Abstract

Abstract: Three-dimensional (3D) super-resolution microscopy, using a biplane detection scheme, termed biplane photo-activated localization microscopy (Biplane FPALM), enables imaging of volumes as thick as whole cells and reveals otherwise unaccessible details of cellular organization [1]. Hence, we attempted to visualize mitochondrial reticulum via the matrix space loaded with mitochondria-addressed Eos, while transfecting cells by lentiviral expression. Our 3D images of single Eos molecules in the matrix space have proven the continuous character of mitochondrial reticulum tubules, i.e., an existence of a highly interconnected major mitochondrial reticulum in insulinoma Ins1E and oxidative-phosphorylation-dependent glutaminolytic hepatoma HepG2 cells [2] (Figure).

   Also, using Eos-conjugate of mitochondrial transcription factor-A (TFAM), we have imaged nucleoids of mitochondrial DNA (mtDNA) in which TFAM represents a major assessor protein. Using PA-CFP2-TFAM and mitochondria-addressed Eos, the first 3D two color super-resolution images were obtained for mitochondrial reticulum together with the distribution of mt nucleoids in it. In intact cells we have found mt nucleoids of a narrow size distribution. The Biplane FPALM technique has proven to be robust and reliable for imaging of mitochondrion and related substructures.

Supported by grants P302/10/0346 (GACR); ME09029 (Czech Ministry of Education); IAA500110701, and M200110902 (Academy of Sciences).) and 1R01GM091791-02 (NIH). Disclosure statement: J.B. declares significant financial interest in Vutara, Inc.

[1] Juette MF, Gould TJ, Lessard MD, Mlodzianoski MJ, Nagpure BS, Bennett BT, Hess ST, Bewersdorf J (2008) 3D sub-100 nm resolution by biplane fluorescence photoactivation localization microscopy. Nat. Methods 5: 527-529.

[2] Mlodzianoski MJ, Schreiner JM, Callahan SP, Smolková K, Dlasková A, Šantorová J, Ježek P, Bewersdorf J (2011) Sample drift correction in 3D fluorescence photoactivation localization microscopy. Opt. Express. 19: 15009-15019. • Keywords: 3D mitochondrial morphology, BiplaneFPALM microscopy, oxidative phosphorylation, mitochondrial DNA

Labels:

Preparation: Intact Cell; Cultured; Primary"Intact Cell; Cultured; Primary" is not in the list (Intact organism, Intact organ, Permeabilized cells, Permeabilized tissue, Homogenate, Isolated mitochondria, SMP, Chloroplasts, Enzyme, Oxidase;biochemical oxidation, ...) of allowed values for the "Preparation" property.