Post-transcriptional Regulation 
of Gene Expression, RNA-Protein Complexes, 
and Nuclear Structures 
Gideon Dreyfuss, Ph.D. — Investigator 
Dr. Dreyfuss is also Professor of Biochemistry and Biophysics at the University of Pennsylvania School 
of Medicine. He received his Ph.D. degree in biological chemistry from Harvard University and his 
postdoctoral research training as a Helen Hay Whitney fellow at the Massachusetts Institute of Technology. 
Prior to his present appointment, he was Professor and Established Investigator of the American Heart 
Association at Northwestern University. 
MESSENGER RNAs (mRNAs) are the func- 
tional translatable intermediates in the 
pathway of gene expression from DNA to pro- 
teins. They are formed in the nuclei of eukaryotic 
cells by extensive and tightly regulated post- 
transcriptional processing of primary RNA poly- 
merase II transcripts. These transcripts are 
termed heterogeneous nuclear RNAs (hnRNAs) , a 
term that describes their size heterogeneity and 
cellular localization. The terms hnRNA and pre- 
mRNA are often used interchangeably, though it 
is possible that only a subset of hnRNAs are actu- 
ally precursors to mRNA and that the rest turn 
over in the nucleus. From the time hnRNAs 
emerge from the transcription complex and as 
long as they remain in the nucleus, they are asso- 
ciated with proteins. 
The collective term for the proteins that bind 
hnRNAs (but are not stable components of other 
classes of ribonucleoprotein complexes, such as 
snRNPs [small nuclear ribonucleoproteins]) is 
hnRNPs. The significance of hnRNPs is that they 
are bound to the hnRNAs and thus influence their 
structure and therefore their fate and processing 
into mRNAs. They are also abundant in the nu- 
cleus and in hnRNA-hnRNP complexes (hnRNP 
complexes) and of interest as major nuclear 
structures. 
Once formed, the mRNAs are transported to the 
cytoplasm via nuclear pores — a process that ap- 
pears to be one of the most important regulatory 
steps in the post-transcriptional pathway of gene 
expression and about which very little is pres- 
ently known. In the cytoplasm, mRNAs are asso- 
ciated with proteins — the mRNPs — and these are 
likely to be involved in the regulation of the 
translation and stability of mRNAs and in their 
cellular localization. Our goal is to understand, 
in molecular detail and cellular architecture, 
how the post-transcriptional portion of the path- 
way of gene expression operates in the cell. To 
that end, we investigate the structure, function, 
and localization of the hnRNPs and mRNPs and 
the RNP complexes. 
We have used photochemical RNA-protein 
crosslinking in intact cells and affinity chromato- 
graphic methods to identify and purify the 
hnRNPs and mRNPs and have produced monoclo- 
nal antibodies to many of them. The monoclonal 
antibodies were used for immunopurification of 
hnRNP complexes from vertebrate and Drosoph- 
ila melanogaster cells and for the characteriza- 
tion of the hnRNPs. Immunopurified hnRNP 
complexes contain large RNA of up to 10,000 
nucleotides and at least 20 major proteins, desig- 
nated A-U, in the range of 34- 1 20 kDa. There are 
also many less abundant hnRNPs, and these ap- 
pear to bind only to specific subsets of hnRNAs. 
The specific arrangement of the proteins on spe- 
cific hnRNAs is probably important in determin- 
ing the structure of the hnRNA and is one of the 
areas on which we concentrate our investigation. 
Related to this issue, we found that several of 
the hnRNPs have RNA-binding specificities. Inter- 
estingly, some of the specificities of the major 
hnRNPs are for sequences important in pre- 
mRNA processing and polyadenylation, and it is 
likely that this binding specificity is directly re- 
lated to a role for these proteins in mRNA 
formation. 
The molecular cloning and sequencing of 
cDNAs for several RNPs made possible the discov- 
ery of a conserved RNA-binding domain (RBD) 
and a ribonucleoprotein consensus sequence 
(RNP-CS). The RNP-CS, Lys/Arg-Gly-Phe/Tyr- 
Gly/Ala-Phe-Val-X-Phe/Tyr, is the most highly 
conserved segment in a generally conserved do- 
main of about 90 amino acids found in many 
RNA-binding proteins of the nucleus and cyto- 
plasm in all eukaryotes examined. Overall, RNP- 
CS proteins have a modular structure reminiscent 
of DNA-binding transcription factors. They fre- 
quently contain several similar but nonidentical 
RBDs, and all contain at least one auxiliary 
domain that is unique to each type of protein 
(e.g., glycine-rich, glutamine-rich, or acidic) and 
that most likely functions in protein-protein 
interactions. 
The hnRNP C1/C2 proteins are abundant, avid 
hnRNA binders. Deletional analysis showed that a 
93-amino acid segment of the human hnRNP C 
protein that contains the RNP-CS is, as predicted, 
117 
