7 research outputs found
Differential expression of FMR1, FXR1 and FXR2 proteins in human brain and testis
Lack of expression of the fragile X mental retardation protein (FMRP)
results in mental retardation and macroorchidism, seen as the major
pathological symptoms in fragile X patients. FMRP is a cytoplasmic
RNA-binding protein which cosediments with the 60S ribosomal subunit.
Recently, two proteins homologous to FMRP were discovered: FXR1 and FXR2.
These novel proteins interact with FMRP and with each other and they are
also associated with the 60S ribosomal subunit. Here, we studied the
expression pattern of the three proteins in brain and testis by
immunohistochemistry. In adult brain, FMR1, FXR1 and FXR2 proteins are
coexpressed in the cytoplasm of specific differentiated neurons only.
However, we observed a different expression pattern in fetal brain as well
as in adult and fetal testis, suggesting independent functions for the
three proteins in those tissues during embryonic development and adult
life
Instability of a (CGG)98 repeat in the Fmr1 promoter
Fragile X syndrome is one of 14 trinucleotide repeat diseases. It arises
due to expansion of a CGG repeat which is present in the 5'-untranslated
region of the FMR1 gene, disruption of which leads to mental retardation.
The mechanisms involved in trinucleotide repeat expansion are poorly
understood and to date, transgenic mouse models containing transgenic
expanded CGG repeats have failed to reproduce the instability seen in
humans. As both cis-acting factors and the genomic context of the CGG
repeat are thought to play a role in expansion, we have now generated a
knock-in mouse Fmr1 gene in which the murine (CGG)8 repeat has been
exchanged with a human (CGG)98 repeat. Unlike other CGG transgenic models,
this model shows moderate CGG repeat instability upon both in maternal and
paternal transmission. This model will now enable us to study the timing
and the mechanism of repeat expansion in mice
The fragile X-related proteins FXR1P and FXR2P contain a functional nucleolar-targeting signal equivalent to the HIV-1 regulatory proteins
Fragile X syndrome is caused by the absence of the fragile X
mental-retardation protein (FMRP). FMRP and the fragile X-related proteins
1 and 2 (FXR1P and FXR2P) form a gene family with functional similarities,
such as RNA binding, polyribosomal association and nucleocytoplasmic
shuttling. In a previous study, we found that FMRP and FXR1P shuttle
between cytoplasm and nucleoplasm, while FXR2P shuttles between cytoplasm
and nucleolus. The nuclear and nucleolar-targeting properties of these
proteins were investigated further. Here, we show that FXR2P contains in
its C-terminal part, a stretch of basic amino acids 'RPQRRNRSRRRRFR' that
resemble the nucleolar-targeting signal (NoS) of the viral protein Rev.
This particular sequence is also present within exon 15 of the FXR1 gene.
This exon undergoes alternative splicing and is therefore only present in
some of the FXR1P isoforms. We investigated the intracellular distribution
of various FXR1P isoforms with (iso-e and iso-f) and without (iso-d) the
potential NoS in transfected COS cells treated with the nuclear export
inhibitor leptomycin-B. Both iso-e and iso-f showed a nucleolar
localization, as observed for FXR2P; iso-d was detected in the
nucleo-plasm outside the nucleoli. Further, when a labelled 16-residue
synthetic peptide corresponding to the NoS of FXR1P was added to human
fibroblast cultures a clear nucleolar signal was observed. Based on these
data we argue that the intranuclear distribution of FXR2P and FXR1P
isoforms is very likely to be mediated by a similar NoS localized in their
C-terminal region. This domain is absent in some FXR1P isoforms as well as
in all FMRP isoforms, suggesting functional differences for this family of
proteins, possibly related to RNA metabolism in different tissues
Different targets for the fragile X-related proteins revealed by their distinct nuclear localizations
Fragile X syndrome is caused by the absence of the fragile X mental
retardation protein (FMRP). FMRP and its structural homologues FXR1P and
FXR2P form a family of RNA-binding proteins (FXR proteins). The three
proteins associate with polyribosomes as cytoplasmic mRNP particles. Here
we show that small amounts of FMRP, FXR1P and FXR2P shuttle between
cytoplasm and nucleus. Mutant FMRP of a severely affected fragile X
patient (FMRPI304N) does not associate with polyribosomes and shuttles
more frequently than normal FMRP, indicating that the association with
polyribosomes regulates the shuttling process. Using leptomycin B we
demonstrate that transport of the FXR proteins out of the nucleus is
mediated by the export receptor exportin1. Finally, inactivation of the
nuclear export signal in two FXR proteins shows that FMRP shuttles between
cytoplasm and nucleoplasm, while FXR2P shuttles between cytoplasm and
nucleolus. Therefore, molecular dissection of the shuttling routes used by
the FXR proteins suggests that they transport different RNAs
Knockout mouse model for Fxr2: a model for mental retardation
Fragile X syndrome is a common form of mental retardation caused by the
absence of the FMR1 protein, FMRP. Fmr1 knockout mice exhibit a phenotype
with some similarities to humans, such as macro-orchidism and behavioral
abnormalities. Two homologs of FMRP have been identified, FXR1P and FXR2P.
These proteins show high sequence similarity, including all functional
domains identified in FMRP, such as RNA binding domains. They have an
overlap in tissue distribution to that of FMRP. Interactions between the
three FXR proteins have also been described. FXR2P shows high expression
in brain and testis, like FMRP. To study the function of FXR2P, we
generated an Fxr2 knockout mouse model. No pathological differences
between knockout and wild-type mice were found in brain or testis. Given
the behavioral phenotype in fragile X patients and the phenotype
previously reported for the Fmr1 knockout mouse, we performed a thorough
evaluation of the Fxr2 knockout phenotype using a behavioral test battery.
Fxr2 knockout mice were hyperactive (i.e. traveled a greater distance,
spent more time moving and moved faster) in the open-field test, impaired
on the rotarod test, had reduced levels of prepulse inhibition, displayed
less contextual conditioned fear, impaired at locating the hidden platform
in the Morris water task and were less sensitive to a heat stimulus.
Interestingly, there are some behavioral phenotypes in Fxr2 knockout mice
which are similar to those observed in Fmr1 knockout mice, but there are
also some different behavioral abnormalities that are only observed in the
Fxr2 mutant mice. The findings implicate a role for Fxr2 in central
nervous system function