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Laboratory
of RNA Biology
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| RESEARCH: Control of alternative pre-mRNA splicing in biology and diseases Alternative
splicing is a common way
of gene regulation that allows the generation
of multiple mRNA and often protein isoforms
from a single gene. Almost all human gene
transcripts are alternatively spliced and some
are known to generate extremely diverse
protein isoforms. This greatly contributes to
the proteomic
complexity, particularly in
neurons and endocrine cells. In
experimental animals, genetic deficiencies in
alternative splicing factors result in
developmental defects or embryonic-lethal
phenotypes. In humans, aberrant
splicing can be resulted from 15-30% of genetic
mutations that cause diseases. It is thus
important to understand alternative splicing
and its regulation in biology and diseases in
the omics era.
For example, in electrically excitable
cells such as neurons, endocrine and muscle
cells, ion channels allow ions in/out of the
cell membranes to generate electrical firing
patterns that are important for cell functions
including memory, hormone secretion and muscle
contraction. These processes are believed to
be critical for higher order phenotype such as
learning, behavior, metabolism and heart
beating. How these processes are finely tuned
during development and in adult life is still
a mystery to researchers. Alternative splicing
provides a unique way to diversify proteins
and may play a critical role here. A number of
studies have indicated that alternative
splicing is involved in adaptive or addictive
changes in neurons by neuronal activity
or alcohol stimulation.
Interestingly, alternative splicing of
some ion channel genes is regulated by
membrane depolarization, the first part of an
action potential, implying a gene expression
change related to the electrophysiological
memory observed in
neurons, or hormone production in response to
experience in life (e.g. exercise
or stress).
However, the molecular basis of the splicing
regulation, particularly after recurrent
stimulation, remains largely unclear.
We have used the STREX (stress
axis-regulated
exon) variant of the Slo1 BK potassium
channel
gene as a model to study how cell signals
regulate the choice of alternative splice
sites in pre-mRNA transcripts. Inclusion of
the STREX exon enhances the calcium
sensitivity of BK channels and likely
modulates cellular electrical properties
related to hearing frequency tuning or
adaptive changes in learning and memory or
tolerance to alcohol. Its regulation by stress
hormones and the calcium/calmodulin-dependent
protein kinase IV (CaMK IV) makes it an
interesting target for dissecting the
components regulating alternative splicing as
well as understanding the impact of splicing
regulation on neuronal electrical properties.
A first step toward this goal was made by
coupling CaMK IV with a pre-mRNA element (CaRRE1)
sufficient to confer CaMK IV response to an
otherwise non-responsive exon. We have
recently identified the splicing factors hnRNP
L and L-Like (LL) as
essential components of the CaMKIV-regulated
splicing of STREX by inhibiting U2AF65 binding
to the upstream 3' splice site. Particularly
for hnRNP L, its Ser513 is
phosphorylated
and essential for the regulation. Other
factors including PTB and
hnRNP K are involved in
the regulation as well. In particular, hnRNP L and LL are required for the differential regulation and protection of the hormone gene expression programs for producing prolactin and growth hormones. Further detailed characterization of this molecular process and its role in homeostatic or adaptive splicing, hormone production and stress response is ongoing .
In recent years, reports by several
labs have also demonstrated that alternative
splicing of the BK
channel and AMPA
receptor genes after chronic
inactivities or of the neurexin
gene upon depolarization/activities of
neurons plays a critical role in the homeostasis
of cellular electrical properties or synaptic
formation. Moreover, the regulation is
mediated by the Ca++/calmodulin-dependent
protein kinase IV (CaMKIV) and its downstream
splicing factors Sam68
or Nova-2,
depending on the target exons. Together, these
observations support a critical role of
depolarization-regulated splicing in hormone
production, neuronal homeostasis or
development.
CaRRE1-like RNA
elements,
even G
tracts, have been found to act
similarly as CaRRE1 in hundreds of human
genes. Together we call these groups of RNA
sequences REPA (regulatory elements
between the polypyrimidine tract and 3' AG. Most of the REPA G
tracts (REPAG) appear to
have emerged in the ancestors of mammals,
likely contributing to the higher abundance of
alternative splicing and proteomic diversity.
We have demonstrated with the REPAG of PRMT5 exon 3
that it contributes to the evolutionary
emergence of a novel splice variant with an
opposite effect on cell cycle. The REPAGs
are widespread and highly enriched in
metazoa and plants, with the highest abundance
in mammals. In particular, they are highly
enriched in the non-coding regions of the
mammalian genomes, where they are critical for
maintaining the integrity or promoting the
diversity of transcriptomes by controlling
allele-dependent splicing. They are also
enriched in the aberrant
3' splice sites of cancer patients
mutated of the 3' splicing factors SF3B1 or
U2AF35. A particular REPAG example for its
importance in genetic diseases is in the CBS gene, mutation of which causes
homocystinuria,
a REPAG prevents its aberrant
splicing.
Other exons studied include those
involved in neuronal function,
human genetic disease or cell growth/apoptosis,
or splicing regulations involving other
signaling pathways including protein acetylation
and methylation.
Analyses of these signal-responsive RNA
elements indicate that they are mostly
mammalian-specific, likely contributing to the
more delicate and dynamic control of
alternative splicing and higher
proteomic complexity.
We hope these studies will
provide molecular details of
splicing changes in cell
physiology, as well as knowledge for cancer
diagnosis/therapy or the correction of aberrant
splicing that causes human genetic
diseases.
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PUBLICATIONS [PubMed]:
18. Nguyen H, Das U, Wang B, Xie
J. The
matrices and constraints of GT/AG splice
sites of more than 1000 species/lineages. Gene,
2018, 660:92-101. ·
*UPDATEd splice
site matrices/constraints from recent
Ensembl releases. **A list
of alternative exons differentially spliced
between male and hermaphrodite
('female') C. elegans ('sex'-specific
alternative splicing in C. elegans)
from this study (Raw RNA-seq reads from 'Kramer et
al,
Genetics 204 (2016) 355-69' in the NCBI SRA database). |
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The REPAG element specifically
among mammals |
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The CaRRE element among
vertebrates (Liu GD, et al., J. Biol.
Chem. 2012, 287:22709–22716) |
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A new group of
introns: REPA element 'inserted' between the
Py and 3'AG and its effects on alternative
splicing |
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*Research
in this lab has been supported by CIHR , NCIC, CBCF,
NSERC, MMSF, MHRC and CFI.
Misce. links:
- Eradication of Polio in China
RNA Meeting in
Winnipeg, June 23-26, 2019 Some RNA/DNA
literature/history:- Why are the snRNAs in splicing called
U1, U2, U4, U5, U6 snRNAs? Here is the original paper
that describes the use of Us in the names for some of
the small nuclear RNAs that are Uridylic acid-rich, by
James L. Hodnett and Harris Busch of the Baylor
University College of Medicine in 1968: Hodnett JL,
Busch H. Isolation and characterization of uridylic
acid-rich 7 S ribonucleic acid of rat liver nuclei. J
Biol Chem. 1968 Dec 25;243(24):6334-42. |PDF|
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- 谁翻译了基 因一词？维 基百科
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College of Medicine |
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University of Manitoba |