Molecular stages of rapid and uniform neuralization of human embryonic stem cells

http://www.nature.com/cdd/journal/v16/n6/abs/cdd200918a.html

Edited by R De Maria

R Bajpai¹, G Coppola², M Kaul^1,3, M Talantova¹, F Cimadamore¹, M Nilbratt¹, D H Geschwind², S A Lipton¹ and A V Terskikh¹

E-mail: terskikh@burnham.org

Abstract

• Insights into early human development are fundamental for our understanding of human biology. Efficient differentiation of human embryonic stem cells (hESCs) into neural precursor cells is critical for future cell-based therapies.
  
• Here, using defined conditions, we characterized a new method for rapid and uniform differentiation of hESCs into committed neural precursor cells (designated C-NPCs).
  
• Dynamic gene expression analysis identified several distinct stages of ESC neuralization and revealed functional modules of coregulated genes and pathways.
  
• The first wave of gene expression changes, likely corresponding to the transition through primitive ectoderm, started at day 3, preceding the formation of columnar neuroepithelial rosettes. The second wave started at day 5, coinciding with the formation of rosettes. The majority of C-NPCs were positive for both anterior and posterior markers of developing neuroepithelium. In culture, C-NPCs became electrophysiologically functional neurons; on transplantation into neonatal mouse brains, C-NPCs integrated into the cortex and olfactory bulb, acquiring appropriate neuronal morphologies and markers. Compared to rosette-NPCs,¹ C-NPCs exhibited limited in vitro expansion capacity and did not express potent oncogenes such as PLAG1 or RSPO3. Concordantly, we never detected tumors or excessive neural proliferation after transplantation of C-NPCs into mouse brains. In conclusion, our study provides a framework for future analysis of molecular signaling during ESC neuralization.

Keywords:

human embryonic stem cells, uniform neural differentiation, expression/coregulation analysis

Abbreviations:

bFGF, basic fibroblast growth factor; BMP, bone morphogenetic protein; C-NPCs, committed neural precursor cells; CNS, central nervous system; EGF, epidermis growth factor; GFAP, glial fibrillary acidic protein; hESC, human embryonic stem cells; mEPSC, miniature excitatory postsynaptic current; SVZ, subventricular zone; WGCNA, weighted gene coexpression network analysis

The Pirh2–keratin 8/18 interaction modulates the cellular distribution of mitochondria and UV-induced apoptosis

http://www.nature.com/cdd/journal/v16/n6/abs/cdd200912a.html

Edited by L Scorrano

S Duan¹, Z Yao¹, Y Zhu¹, G Wang¹, D Hou¹, L Wen¹ and M Wu
E-mail: wumian@ustc.edu.cn

Abstract

• Intermediate filaments (IFs) provide crucial structural support in higher eukaryotic cells. Accumulating evidences show that IFs also participate in various cellular activities including stress responses, cell growth, cell death and cell migration through dynamic interactions with various non-structure proteins.
  
• Here we report the identification of Pirh2, a RING-H2-type ubiquitin E3 ligase, as a novel binding partner of the cytoplasmic IF proteins keratin 8/18 (K8/18).
  
• Phosphorylation of either Pirh2 or K8/18 affects their association. Although Pirh2 was not found to influence the stability of K8/18, it displayed an unexpected role in regulating the organization of the network of K8/18 keratin filaments.
  
• Disruption of Pirh2–K8/K18 interaction by either UV irradiation or knockdown with Pirh2 or K18 led to the aggregation of K8/18 keratin filaments.
  
• It further induced mitochondrial redistribution, and this process is likely through a microtubule-mediated pathway.
  
• The abnormal localization of mitochondria in Pirh2-knockdown cells may partially account for its increased cell sensitivity to UV-induced apoptosis, probably through enhancing the release of pro-apoptotic proteins, such as cytochrome c and Smac/DIABLO to the cytosol. Overall, our data reveal the novel role of the Pirh2–K8/18 complex in governing the distribution of mitochondria.

Keywords:

Pirh2, keratin 8/18, mitochondria, apoptosis

Abbreviations:

IFs, intermediate filaments; TRADD, TNF receptor type 1-associated death domain protein; JNK, c-Jun N-terminal kinase; TNF, tumor necrosis factor