Presence and phosphorylation of transcription factors in developing dendrites

Presence and phosphorylation of transcription factors in developing dendrites

Departments of ^*Pharmacology, ^†Physiology, ^§Pathology, and ^¶Psychiatry, University of Pennsylvania School of Medicine, Philadelphia, PA 19104; and ^‡National Human Genome Research Institute, National Institutes of Health, Bethesda, MD 20892

Edited by Bert O’Malley, Baylor College of Medicine, Houston, TX, and approved December 19, 1997 (received for review November 14, 1997)

Abstract

In screening amplified poly(A) mRNA from hippocampal dendrites and growth cones in culture to determine candidates for local translation, we found that select transcription factor mRNAs were present. We hypothesized that synthesis of transcription factor proteins within dendrites would provide a direct signaling pathway between the distal dendrite and the nucleus resulting in modulation of gene expression important for neuronal differentiation. To evaluate this possibility, radiolabeled amplified antisense RNA was used to probe slot blots of transcription factor cDNAs as well as arrayed blots of zinc finger transcription factors. The mRNAs encoding the cAMP response element binding protein (CREB), zif 268, and one putative transcription factor were detected. We expanded upon these results showing that CREB protein is present in dendrites, that translation of CREB mRNA in isolated dendrites is feasible and that CREB protein found in dendrites can interact with the cis-acting cyclic AMP reponse element DNA sequence by using an in situ Southwestern assay. Further, CREB protein in dendrites is not transported to this site from the cell body because fluorescently tagged CREB microperfused into the soma did not diffuse into the dendrites. In addition, CREB protein microperfused into dendrites was rapidly transported to the nucleus, its likely site of bioactivity. Lastly, by using the isolated dendrite system we show that phosphorylation of Ser-133 on CREB protein can occur in isolated dendrites independent of the nucleus. These data provide a regulatory pathway in which transcription factors synthesized and posttranslationally modified in dendrites directly alter gene expression bypassing the integration of signal transduction pathways that converge on the nucleus.

Footnotes

↵ ¶ To whom reprint requests should be addressed. e-mail: eberwine{at}mscf.med.upenn.edu.
This paper was submitted directly (Track II) to the Proceedings Office.
Abbreviations: aRNA, amplified antisense RNA; CRE, cyclic AMP response element; CREB, CRE binding protein; pBS, pBluescript plasmid cDNA; DHPG, 3,5-dihydroxyphenylglycine.