Expression of chondroitin sulfotransferases in neural progenitor cells : impacts on cell motility

Abstract

The specific order by which functional neural circuits are established relies on the timely migration of newly specified neurons to target sites where they complete the differentiation program and form synapses. Chondroitin sulfate (CS) proteoglycans are well-positioned in the extracellular matrix of developing neural tissue for roles in cell proliferation, migration, growth cone guidance and axon fasciculation. In this study, we hypothesize that the newly born neurons tune into chemotactic signals in the environment by modulating the sulfation pattern of perineuronal chondroitins. We chose to study the migration of such neurons in hindbrain explants (E11.5+ 1 day-in-vitro, Sprague Dawley rats) using time lapse microscopy. In control cultures, neurons advanced along exiting neurites that radiated from the explant. In test cultures treated with chondroitinase ABC, neurons lost directional migration but retained motility; immunocytochemistry confirmed loss of the CS-56 epitope in the chondroitinase ABC-treated cultures. Re-examination of the control culture revealed that the migrating neurons close to the explant core engaged in more active forward movements compared to those in the periphery (near the distal end of neurites). The regrowth of CS-56 epitope positive CS moieties not only rescued the somal migration previously halted by ChABC treatment but also enhanced the advancement of migrating neurons. Fluorescence in situ hybridization for mRNAs of CS-sulfotransferases revealed that the migratory neurons largely expressed chondroitin-6-sulfotransferase-1, chondroitin-4-sulfotransferase-2, N-acetylgalactosamine 4-sulfate 6-O-sulfotransferaseand uronal-2-sulfotransferase whereas those remaining in the explant core signals largely expressed chondroitin-4-sulfotransferase-1. CS-56 epitopes were detectable in nestin-and glial fibrillary acidic protein-positive-neural precursors derived from human bone marrow stromal cells as well. Semi-quantitative RT-PCR and fluorescence in situ hybridization for mRNAs of CS-sulfotransferases revealed that the mRNA expression of chondroitin-4-sulfotransferase-2remained high while chondroitin-4-sulfotransferase-1 and chondroitin-6-sulfotransferase-1 showed sudden changes during the days of active progenitor migration as shown in time-lapse video recordings. Our results suggest a new mechanism by which tuning sulfation of perineuronal chondroitins confers sensitivity of early hindbrain motor neuron progenitors to directional cues in the extracellular environment and that the sulfation patterns of cell surface CS on neurons are modified to overcome the inhibition imposed by the environmental CS deposited. Our preliminary data also suggest the roles of 4-and 6-O-sulfated CS by modified by chondroitin-4-sulfotransferase-2and chondroitin-6-sulfotransferase-1 in migrating neural precursors.