Laboratory of RNA Biology |
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RESEARCH:
We
are
interested
in
understanding how the different fragments of a gene are
selectively combined (spliced) to produce multiple
products in biology and diseases. This process, called
pre-mRNA splicing, has profound implications in the
evolution and functional complexity of genes/gene
products. Its disruption is
a cause of human genetic diseases by
many mutations and also of the aberrant splicing in
cancers. Our past research has focused on the molecular
basis of adaptive splicing, likely involved in
cell adaptation, neuron homeostasis and neurological
diseases. This has also allowed us to discover and
extend the scope to a novel class of introns/splice
sites (REPA-harbouring) contributing to
the evolution of alternative splicing, gene product
diversity or to the prevention of aberrant splicing in
human health/diseases. Particularly
welcome to join us are students with backgrounds in
biochemistry, genetics, computer science or statistics.
Control of alternative pre-mRNA splicing in biology
and diseases
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 Slo1BK 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.(Fig. 1) 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, Fig. 3). 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.
They are also enriched in the aberrant
3' splice sites of cancer patients mutated of
the 3' splicing factors SF3B1 or U2AF35. Moreover, a
REPAG prevents aberrant
splicing of the CBS
gene, mutation of which causes
the human genetic disease
homocystinuria.
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.
Ogunsola S, Liu L, Das U, Xie J. 5-Aza-Cytidine Enhances Terminal Polyadenylation Site Usage for Full-Length Transcripts in Cells. Genes to Cells (2024) In revision. bioRxiv: 2024.2002.2022.581641.
2.
Liu
L,
Nguyen H, Das U, Ogunsola S, Yu J, Lei L, Kung M, Pejhan
S, Rastegar M, Xie J: Epigenetic Control of Adaptive or
Homeostatic Splicing During Interval-Training
Activities. Nucleic Acids Res. (2024) In
revision. bioRxiv 2024:2024.02.23.581772. https://doi.org/10.1101/2024.02.23.581772
3.
Joyce W, He K,
Zhang M, Ogunsola S, Wu X, Joseph KT, Bogomolny D, Yu W,
Springer MS, Xie J, Signore AV, Campbell KL: Parallel
genetic excisions of the cardiac troponin I N-terminal
extension in tachycardic mammals. bioRxiv
2024:2023.05.19.541292. https://doi.org/10.1101/2023.05.19.541292
4.
Xie, J.,
Wang, L. & Lin, RJ. Variations of intronic
branchpoint motif: identification and functional
implications in splicing and disease. Commun Biol 6,
1142 (2023). https://doi.org/10.1038/s42003-023-05513-7.
5.
Xie J and Friedman R,
Editorial: Evolution in Neurogenomics. Front. Genet. (2023)14:1220750.
doi:
10.3389/fgene.2023.1220750.
6.
Liu L, Das U, Ogunsola S, Xie
J. Transcriptome-Wide Detection of Intron/Exon
Definition in the Endogenous Pre-mRNA Transcripts of
Mammalian Cells and its Regulation by Depolarization, Int.
J. Mol. Sci. (2022) 23, 10157. https://
doi.org/10.3390/ijms231710157
7.
Tian L, Xie X, Das U, Chen Y,
Sun Y, Liu F, Lu H, Peng N, Zhu Y, Gu X, Deng H, Xie J,
Zhao X. Forming cytoplasmic stress granules PURα
suppresses mRNA translation initiation of IGFBP3 to
promote esophageal squamous cell carcinoma progression,
Oncogene, 41, 4336–4348 (2022), https://www.nature.com/articles/s41388-022-02426-3.
8.
Ling Liu,
Jinghua Feng, Julian Polimeni, Manli Zhang, Hai Nguyen,
Urmi Das, Xu Zhang, Harminder Singh, Xiao-Jian Yao,
Etienne Leygue, Sam K.P. Kung, and Xie J. Characterization of cell free plasma
methyl-DNA from xenografted tumours to guide the
selection of diagnostic markers for early-stage cancers. Frontiers
in Oncology,
11:615821 (2021) Feb. 5. DOI: 10.3389/fonc.2021.615821.
9.
Xie J,
Weiskirchen R. What does the 'AKT' stand for in the
name 'AKT kinase'?: some historical comments. Frontiers
in Oncology, 10:1329. June, 2020, https://www.frontiersin.org/articles/10.3389/fonc.2020.01329/full.
10.
Nguyen H,
Das U, Xie J. Genome-wide evolution of wobble
base-pairing nucleotides of branchpoint motifs with
increasing organismal complexity. RNA Biology, 17:3, 311-324, Dec. 2019, http://dx.doi.org/10.1080/15476286.2019.1697548.
11.
Nguyen H,
Xie J. Widespread
separation of the polypyrimidine tract from 3' AG by G
tracts in association with alternative exons in metazoa
and plants. Frontiers in Genetics, 9:741,
published: 14 January 2019, doi:
10.3389/fgene.2018.00741. |PDF|
12.
Das U,
Nguyen H, Xie J. Transcriptome protection by the expanded
family of hnRNPs. RNA Biology, 16:2, 155-159, 2018 Dec 30. PMID:
30596342 DOI: 10.1080/15476286.2018.1564617.
13.
Lei L,
Cao W, Liu L, Das U, Wu Y, Liu G, Sohail M, Chen Y, Xie
J. Multi-level
differential
control of hormone gene expression programs by hnRNP L
and LL in pituitary cells. Mol.
Cell.
Biol., 2018 May 29;38(12):e00651-17.(MCB Most-Read'04-05'18). |PDF|
14.
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.
------ *UPDATE 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).
Fig. 1. Molecular basis of the depolarization-regulated alternative splicing of the STREX exon of the Slo1 gene.
Fig. 2. Importance of proper splicing control in hormone production: aberrant splice variant of prolactin (right) due to the loss of hnRNP L (image by Lei Lei).Fig. 3. A new group of introns: REPA element 'inserted' between the Py and 3'AG and its effects on alternative splicing.