Biology subjects

Doddihal, V.

Publications and source records attributed to Doddihal, V..

4 recordsLinked to original sources

bioRxiv2022

A PAK kinase family member and the Hippo/Yorkie pathway modulate WNT signaling to functionally integrate body axes during regeneration

Successful regeneration of missing tissues requires seamless integration of positional information along the body axes. Planarians, which regenerate from almost any injury, use conserved, developmentally important signaling pathways to pattern the body axes. However, the molecular mechanisms which facilitate crosstalk between these signaling pathways to integrate positional information remain poorly understood. Here, we report a p21-activated kinase (smed-pak1) which functionally integrates the anterior-posterior (AP) and the medio-lateral (ML) axes. pak1 inhibits WNT/{beta}-catenin signaling along the AP axis and, functions synergistically with the {beta}-catenin-independent WNT signaling of the ML axis. Furthermore, this functional integration is dependent on warts and merlin - the components of the Hippo/Yorkie (YKI) pathway. Hippo/YKI pathway is a critical regulator of body size in flies and mice, but our data suggest the pathway is required to pattern body axes in planarians. Our study provides a signaling network integrating positional information which can mediate coordinated growth and patterning during planarian regeneration.

developmental biology↗

bioRxiv2021

A powerful and versatile new fixation protocol for immunohistology and in situ hybridization that preserves delicate tissues in planaria

Whole-mount in situ hybridization (WISH) is a powerful and widely used technique to visualize the expression pattern of genes in different biological systems. Here we describe a new protocol for ISH and immunostaining in the planarian Schmidtea mediterranea. The new Nitric Acid/Formic Acid (NAFA) protocol is compatible with both assays and prevents degradation of the epidermis or blastema. Instead of proteinase K digestion, formic acid treatment is used to permeabilize tissues and preserve antigen epitopes. We show that the NAFA protocol successfully permits development of chromogenic and fluorescent signals in situ, while preserving the anatomy of the animal. Further, the immunostaining of different proteins was compatible with the NAFA protocol following fluorescent in situ hybridization. Finally, we demonstrate with high resolution confocal imaging that the regeneration blastema is preserved when using the new method. This new NAFA protocol will be a valuable technique to study the process of wounding response and regeneration.

developmental biology↗

bioRxiv2020

Decellularization enables functional analysis of ECM remodeling in planarian regeneration

The extracellular matrix (ECM) is a three-dimensional network of macromolecules that provides a microenvironment capable of supporting and regulating cell functions. However, only a few research organisms are available for the systematic dissection of the composition and functions of the ECM, particularly during regeneration. We utilized a free-living flatworm Schmidtea mediterranea to develop an integrative approach consisting of decellularization, proteomics, and RNA-interference (RNAi) to characterize and investigate ECM functions during tissue homeostasis and regeneration. High-quality ECM was isolated from planarians, and its matrisome profile was characterized by LC-MS/MS. The functions of identified ECM components were interrogated using RNAi. Using this approach, we discovered that heparan sulfate proteoglycan and kyphoscoliosis peptidase are essential for both tissue homeostasis and regeneration. Our strategy provides a robust experimental approach for identifying novel ECM components involved in regeneration that might not be discovered bioinformatically.

developmental biology↗

bioRxiv2020

The Planarian Anatomy Ontology: A resource to connect data within and across experimental platforms

As the planarian Schmidtea mediterranea (Smed) gains popularity as a research organism, the need for standard anatomical nomenclature is increasingly apparent. A controlled vocabulary streamlines data annotation, improves data organization, and enhances cross-platform and cross-species searchability. We created the Planarian Anatomy Ontology (PLANA), an extendable framework of defined Smed anatomical terms organized using relationships. The most current version contains over 800 terms that describe Smed anatomy from subcellular to system-level across all life cycle stages, in intact animals, and regenerating body fragments. Terms from other anatomy ontologies were imported into PLANA to promote ontology interoperability and comparative anatomy studies. To demonstrate the utility of PLANA for data curation, we created web-based resources for planarian embryogenesis, including a staging series and molecular fate mapping atlas, as well as a searchable Planarian Anatomy Gene Expression database, which integrates a variety of published gene expression data and allows retrieval of information of all published sequences associated with specific planarian anatomical regions. Finally, we report methods for continued curation of PLANA, providing a path for expansion and evolution of this community resource. Summary StatementWe report construction of an anatomy ontology for an emerging research organism and show its use to curate and mine data across multiple experimental platforms.

developmental biology↗