|Globin mRNA Stability
Decay rates of individual mRNAs can differ by over 100-fold; with half-lives as short as 15 minutes and as long as several days. This dramatic span of stabilities can have a powerful impact on patterns of gene expression. In addition, a decay rate for an mRNA need not be static and can be regulated as a consequence of feedback mechanisms, the presence of external factors within the cellular environment, a particular stage of differentiation or of the cell-cycle, or viral infection. Our laboratory has a long-standing interest in understanding the mechanistic and physical basis for these differences in mRNA stability and the controls that they exert over gene expression during cellular differentiation and organismal development.
An interesting example highlighting the importance of mRNA stability during cellular differentiation is represented by the globin mRNAs. Terminally differentiating human erythroid cells are characterized by an accumulation of high levels of a - and b -globin proteins. This high level of gene expression is dependent on an unusually long half-life of the a - and b -globin mRNAs (in the order of 24-60 h). This stability facilitates the accumulation of these transcripts in transcriptionally-active erythroid precursors, and ensures that they remain at high levels in subsequent, transcriptionally silent stages of terminal erythroid differentiation. a - and b -globin mRNAs represent more than 95% of total mRNA in the more differentiated red blood cells due to a selective stability of globin mRNAs and degradation of non-globin mRNAs.
A naturally occurring a -globin mutant, a -Constant Spring, clearly demonstrate the
importance of mRNA stability in a -globin gene expression. The a CS mutation is a base change in the normal
stop codon of the a 2-globin reading frame promoting read-through
into the 3'-UTR for an additional 31 codons. This read-through results in destabilization of the
a -globin mRNA, loss of >95% of a -globin gene expression from the affected
locus, and resultant clinical disease ( a -Constant Spring thalassemia). These studies
lead to the hypothesis that the ribosomal read-through caused disruption of a protein complex that is
normally present on the a
-globin 3'-UTR and which confers
high stability of this transcript in erythroid cells. Indeed, the stability of a -globin mRNA in vivo was shown to be
paralleled by the ability of its 3'-UTR to assemble an RNP complex (termed the ' a -complex') in vitro . Mutations that
disrupt a -complex formation in vitro decrease
a -globin mRNA stability in vivo . The
cis -acting sequences necessary for a -complex assembly comprise three
pyrimidine-rich regions. Two closely related (80% identical) proteins a CP-1 and -2 have been purified by virtue of
RNA affinity chromatography using a -globin 3'-UTR sequences, and the
corresponding genes subsequently cloned.
(See Figure 2.) These proteins
each contain three copies of the KH domains found in a number of RNA binding proteins. a CP-1 and -2 are both incorporated into the
a -complex in vitro. We
have demonstrated that the a
-complex is a binary complex
containing a single molecule of the a -CP protein bound to the pyrimidine-rich
cis element. Moreover, we have recently shown that the a -complex can be maintained on actively
translating mRNAs and that aCP can stabilize the a -globin mRNA even when artificially tethered
to the 3'UTR. These findings lead us to a model of mRNA stabilization in which the a CP works via a set of secondary
protein-protein interactions. We are now in the process of defining the full spectrum of mRNAs that
are targeted by a CP and defining in each case the function of
these complexes and their modes of action. On particularly informative approach that we are taking is
to probe the function of the nuclear and cytoplasmic a CP proteins using a set of specifically
designed small RNAs that can effectively 'decoy' a CP from its normal a -globin mRNA target. (See Figure 3.)
||Xinjun Ji, Ph.D. (recipient of a Cooley's Anemia
Foundation Research Fellowship)
||Shelly Waggoner, Ph.D. (Instructor in Genetics
and recipient of an NIH K01 Award)
||Jian Kong, Ph.D.
||Marina Sumaroka, Ph.D.
Primary Papers (1995 - present)
- Wang, X., Kiledjian, M., Weiss, I.M., and Liebhaber, S.A. 1995. Detection and characterization of a 3' untranslated region ribonucleoprotein complex associated with human a-globin mRNA stability. Mol. Cell. Biol. 15:1769-1777.
- Weiss, I.M., and Liebhaber, S.A. 1995. Erythroid-cell specific mRNA stability elements in the a 2-globin 3' untranslated region. Mol. Cell. Biol. 15:2457-2465.
- Kiledjian, M., Wang, W., and Liebhaber , S.A. 1995. Identification of two KH domain proteins in the a -globin mRNP stability complex. EMBO 14:4357-4364.
- Russell, J.E., and Liebhaber, S.A. 1996. The stability of human ß-globin mRNA is dependent upon structural determinants positioned within its 3' untranslated region. Blood 87:5314-5324.
- Wang, X., and Liebhaber, S.A. 1996. Complementary change in Cis determinants and trans factors in the evolution of an mRNP stability complex. EMBO 15:5040-5051.
- Morales, J., Russell, J.E., and Liebhaber, S.A. 1997. Destabilization of human a -globin mRNA by anti-termination is controlled during erythroid differentiation and is parallel by phased shortening of the poly(A) tail. J. Biol. Chem. 272:6607-6613.
- Holcik, M., and Liebhaber, S.A. 1997. RNP complexes sharing cis and trans components assemble at the 3'UTR of four highly stable mRNAs. PNAS 94:2410-2414.
- Stefanovic, B., Hellerbrand, C., Holcik, M., Briendl, M., Liebhaber, S.A, and Brenner, D.A. 1997. Posttranscriptional regulation of collagen alpha (1) mRNA in hepatic stellate cells. Mol. Cell. Biol. 17:5201-5209.
- Russell, J.E., Morales, J., Makayev, A., and Liebhaber, S.A. 1998. Sequence divergence in the 3' untranslated regions of human z - and a -globin mRNAs mediates a difference in their stabilities and contributes to efficient a -to- z gene developmental switching . Mol. Cell. Biol. 18:2173-2183.
- Russell, J.E. and Liebhaber, S.A. 1998. Reversal of a and b thalassemia in adult mice by expression of human embryonic globins. Blood 92:3057-3063.
- Chkheidze, A.N., Lyakhov, D.L., Makeyev, A.V., Morales, J., Kong, J., and Liebhaber , S.A. 1999. Assembly of the a -Globin mRNA stability complex reflects binary interaction between the pyrimidine-rich 3'UTR determinant and the polyC binding protein, a CP. Mol. Cell. Biol., 19:4572-4581.
- Makeyev, A.V., Chkheidze, A.N., and Liebhaber , S.A. 1999. A set of highly conserved RNA-binding proteins, a CP-1 and a CP-2, implicated in mRNA stabilization, are coexpressed from an intronless gene and its intron-containing paralog. J. Biol. Chem, 274:24849-24857.
- Makeyev, A.V. and Liebhaber , S.A. 2000. Identification of two novel mammalian genes establishes a subfamily of KH-domain RNA-binding proteins. Genomics, 67:301-316.
- Thisted, T., Lyakhov, D.L., and Liebhaber , S.A. 2001. Optimized RNA targets of two closely-related triple KH-domain proteins, hnRNP K and aCP-KL, suggest distinct modes of RNA recognition. JBC 276:17484-17496.
- Makeyev, AV, Eastmond, DL, and Liebhaber, SA., 2002. Targeting a KH-domain protein with RNA decoys. RNA, 8:1160-1173
- Ji, X., Kong, J., and Liebhaber, SA 2003. In Vivo Association of the Stability Control Protein, a CP, with Actively Translating mRNAs. Mol Cell Biol 23, 899-907
- Kong, J., Ji, X., and Liebhaber, SA 2003. The KH-Domain protein? a CP has a direct role in mRNA stabilization independent of its cognate binding site. Mol Cell Biol 23, 1125-1134
- Waggoner, S.A. and Liebhaber , S.A. 2003. Identification of mRNAs associated with a CP-2 mRNP complexes. Mol Cell Biol. 23,7055-7067
- Chkheidze A. N. and Liebhaber S. A. 2003. A Novel Set of Nuclear Localization Signals Determine Distributions of the a CP RNA-binding Proteins. Mol Cell Biol. 23,8405-8415. Download
- Inácio Â, Silva AL , Pinto J , Ji X, Morgado A, Almeida F, Faustino P , Lavinha J, Liebhaber SA and Romão L. 2004. Nonsense Mutations in Close Proximity to the Initiation Codon Fail to Trigger Full Nonsense-Mediated mRNA Decay. J. Biol. Chem. In Press.
- Kong Jian, Sumaroka Marina, Eastmond Dawn, and Stephen A. Liebhaber. 2006. Shared Stabilization Functions of Pyrimidine-rich Determinants in the Erythroid 15-Lipoxygenase and ±-Globin mRNAs. Mol Cell Biol. 26:5603-5614. Download
Reviews (1995 - present)
- Liebhaber , S.A. , Wang, X., Kiledjian, M., and Weiss, I.M. 1995. Cis and trans control of globin mRNA stability. In: Proceedings of the 9th Conference on Hemoglobin Switching, 31:375-389.
- Russell, J.E., Morales, J., and Liebhaber, S.A. 1997. The role of mRNA stability in the control of globin gene expression. In: Progress in Nucleic Acid Research and Molecular Biology, 57:249-287.
- Holcik, M., and Liebhaber, S.A. 1997. Analysis of mRNP complexes asembled in vitro. In: mRNA Formation and Function pp. 196-208 (ed. Joel Richter). Academic Press, Publishers.
- Liebhaber , S.A. 1997. mRNA stability and the control of gene expression. In: Nucleic Acids Research Symposia Series No. 36 pp. 29-32. Oxford University Press, Publishers.
- Liebhaber, S.A., and Russell, J.E. 1998. Expression and developmental control of the human a -globin gene cluster. In: Seventh Cooley's Anemia Symposium. Annals of the New York Academy of Sciences, Publishers. Vol. 850:386-390.
- Liebhaber , S.A. 1999. Book Review: Eucaryotic mRNA Processing. Frontiers in Molecular Biology. In Quarterly Review of Biology. Vol. 74:215-216.
- Liebhaber , S.A. 2000. Regulation of a -Globin mRNA Stability. In mRNA Binding Proteins. In Press
- Makayev, A.V. and Liebhaber , S.A. 2002. A Review. Poly C binding proteins: A Multiplicity of Functions and a Search for Mechanisms. RNA. 8:265-278. Download
- Waggoner S., and Liebhaber , S.A. 2003. Regulation of a-Globin mRNA Stability. Experimental Biology and Medicine. In Press. Download
- Liebhaber S.A. 2002. Pathways of Mammalian Messenger RNA Degradation. In Gene Transfer and Expression in Mammalian Cells. (part of the series New Comprehensive Biochemistry). Elseivier Science Press. In Preparation.
- Liebhaber S.A. 2002. mRNA Stability and the Control of Gene Expression. In Encyclopedia of the Human Genome. (Nature Publishing Group).
- Inacio A., and Liebhaber S.A. 2003. Pathways of Mammalian Messenger RNA Degradation. In Gene Transfer and Expression in Mammalian Cells. (part of the series New Comprehensive Biochemistry). Elseivier Science Press. Chapter 17. (pp 495-513).
- Waggoner S., and Liebhaber , S.A. 2003. Regulation of a -Globin mRNA Stability. In Experimental Biology and Medicine. 228:387-95
Last Updated: June 17, 2005