Cryosectioning and immunofluorescence of C. elegans reveals endogenous polyphosphate in intestinal endo-lysosomal organelles.
Ellen Quarles, Lauren Petreanu, Anjali Narain, Aanchal Jain, Akash Rai, Joyful Wang, Bryndon Oleson, Ursula Jakob
Author Information
Ellen Quarles: University of Michigan, Molecular, Cellular, and Developmental Biology Department, Ann Arbor, MI, USA. Electronic address: equarles@gmail.com.
Lauren Petreanu: University of Michigan, Molecular, Cellular, and Developmental Biology Department, Ann Arbor, MI, USA.
Anjali Narain: University of Michigan, Molecular, Cellular, and Developmental Biology Department, Ann Arbor, MI, USA.
Aanchal Jain: University of Michigan, Molecular, Cellular, and Developmental Biology Department, Ann Arbor, MI, USA.
Akash Rai: University of Michigan, Molecular, Cellular, and Developmental Biology Department, Ann Arbor, MI, USA.
Joyful Wang: University of Michigan, Molecular, Cellular, and Developmental Biology Department, Ann Arbor, MI, USA.
Bryndon Oleson: University of Michigan, Molecular, Cellular, and Developmental Biology Department, Ann Arbor, MI, USA.
Ursula Jakob: University of Michigan, Molecular, Cellular, and Developmental Biology Department, Ann Arbor, MI, USA.
Polyphosphate (polyP) is a ubiquitous polyanion present throughout the tree of life. While polyP's widely varied functions have been interrogated in single-celled organisms, little is known about the cellular distribution and function of polyP in multicellular organisms. To study polyP in metazoans, we developed the nematode Caenorhabditis elegans as a model system. We designed a high-throughput, longitudinal-orientation cryosectioning method that allowed us to scrutinize the intracellular localization of polyP in fixed C. elegans using fluorescent polyP probes and co-immunostaining targeting appropriate marker proteins. We discovered that the vast majority of polyP is localized within the endo-lysosomal compartments of the intestinal cells and is highly sensitive toward the disruption of endo-lysosomal compartment generation and food availability. This study lays the groundwork for further mechanistic research of polyPs in multicellular organisms and provides a reliable method for immunostaining hundreds of fixed worms in a single experiment.
