EIF4E Mouse

What is the structure and molecular weight of mouse eIF4E?

Mouse eIF4E is a cap-binding protein with a molecular weight of approximately 25 kDa as detected by Western blot analysis . The human eIF4E protein (which shares high homology with mouse) spans from Met1 to Val217 . Its primary function is recognizing the 5' cap structure of mRNAs to initiate translation. When conducting Western blot analysis, eIF4E can be detected using specific antibodies at concentrations of 0.1-0.5 μg/mL under reducing conditions .

How do I detect eIF4E expression in mouse tissues and cell lines?

Several validated methods exist for detecting eIF4E in mouse samples:

Immunofluorescence/Immunocytochemistry:

• Use specific antibodies at ~10 μg/mL concentration

• Incubate with samples for approximately 3 hours at room temperature

• Counterstain with DAPI to visualize nuclei

• Use appropriate fluorophore-conjugated secondary antibodies

Western Blot Analysis:

• Prepare lysates from mouse cell lines (MCF-7, Balb/3T3, PC-12)

• Use PVDF membrane and probe with anti-eIF4E antibodies

• Include appropriate loading controls (e.g., GAPDH)

• Use knockout cell lines as negative controls to verify specificity

Is eIF4E essential for normal mouse development?

Surprisingly, while eIF4E is considered critical for translation, Eif4e haploinsufficient mice (Eif4e+/-) with 50% reduction in eIF4E levels are viable and develop normally . These mice:

• Are born at normal Mendelian ratios

• Display normal body weight and survival

• Show no impairment in global protein synthesis

• Have normal cap-dependent and IRES-mediated translation

This finding suggests that eIF4E exists in excess of what is required for normal development and physiology, with the threshold for normal function being below 50% of wild-type levels.

What specialized translational programs are regulated by eIF4E in mice?

Gene set enrichment analysis has revealed that eIF4E regulates distinct classes of mRNAs, particularly under conditions of cellular transformation:

mRNAs with structured 5'UTRs are particularly dependent on eIF4E levels for efficient translation . Network interactome analysis shows that specific functional classes of genes require sufficient eIF4E dose for translation during cellular transformation, revealing an "oncogenic translation program" .

How does eIF4E haploinsufficiency affect oncogenic transformation in mouse models?

While 50% reduction in eIF4E is compatible with normal development, it significantly impedes cellular transformation:

• Eif4e+/- mice are remarkably resistant to transformation

• Cancer cells hijack the excess eIF4E present beyond what's needed for normal development

• eIF4E dose is essential for translating mRNAs regulating reactive oxygen species that fuel transformation

• eIF4E levels regulate a translational program supporting tumorigenesis

This suggests a therapeutic window where targeting eIF4E could impair cancer growth while sparing normal physiological processes.

What is the relationship between eIF4E and neurological functions in mouse models?

eIF4E plays crucial roles in neurological function, particularly in the context of Fragile X Syndrome (FXS):

• eIF4E interacts with FMRP (Fragile X Mental Retardation Protein) via CYFIP1 (cytoplasmic FMRP-interacting protein 1)

• CYFIP1 shuttles between the FMRP-eIF4E complex and the Rac1-Wave regulatory complex, connecting translational regulation to actin dynamics and dendritic spine morphology

• Inhibition of eIF4E-eIF4G interactions with 4EGI-1 normalizes hippocampus-dependent memory deficits in FXS model mice

• 4EGI-1 treatment reverses defects in context discrimination learning

These findings highlight eIF4E's role beyond basic translation in regulating complex neurological functions.

How does mGluR-LTD differ between wild-type and Fragile X Syndrome mouse models, and how is it affected by eIF4E modulation?

mGluR-LTD (metabotropic glutamate receptor-dependent long-term depression) exhibits distinct characteristics in wild-type versus FXS mice:

These results indicate that mGluR-LTD in FXS mice occurs through mechanisms that are dependent on eIF4E-eIF4G interactions but independent of general protein synthesis . This suggests that targeting eIF4E specifically may be more effective than general protein synthesis inhibitors for treating FXS-related synaptic abnormalities.

What is the role of eIF4E in controlling actin dynamics and dendritic spine morphology?

eIF4E is involved in a complex regulatory network affecting neuronal structure:

• CYFIP1 forms a bridge between translational regulation (via eIF4E) and actin cytoskeleton regulation (via Rac1)

• In FXS model mice, there is dysregulation of both CYFIP1/eIF4E and CYFIP1/Rac1 interactions

• 4EGI-1 treatment normalizes enhanced Rac1-PAK-cofilin signaling and altered actin dynamics in FXS mice

• 4EGI-1 also normalizes the increased density of dendritic spines in the hippocampus of FXS model mice

These findings suggest that an imbalance between protein synthesis and actin dynamics contributes to pathophysiology in FXS, and that targeting eIF4E can restore this balance.

What genetic tools are available for studying eIF4E function in mice?

Several genetic models have been developed:

• Complete Eif4e knockout mouse models

• Eif4e haploinsufficient mice (Eif4e+/-) for studying the effects of reduced eIF4E levels

• Transgenic mice expressing bicistronic luciferase reporters for simultaneous measurement of cap-dependent and IRES-dependent translation

• Cell-specific conditional knockout models for tissue-specific studies

These tools allow researchers to analyze eIF4E function in various physiological and pathological contexts.

How can I pharmacologically modulate eIF4E function in mouse models?

4EGI-1 is a well-characterized inhibitor that blocks eIF4E-eIF4G interactions:

• For in vivo studies: Infuse 4EGI-1 (100 μM) into the lateral ventricle of mice

• For ex vivo studies: Apply 4EGI-1 (100 μM) to hippocampal slices

• Effects can be observed within 1 hour of administration

• 4EGI-1 does not alter baseline synaptic transmission in either wild-type or FXS model mice

Using pharmacological modulators in combination with genetic models provides complementary approaches to understand eIF4E function in various contexts.

What approaches can be used to study eIF4E-dependent translation in mouse models?

Several techniques have proven valuable:

• Ribosome profiling and genome-wide translational profiling to identify eIF4E-sensitive mRNAs

• Western blotting with eIF4E knockout cell lines as negative controls

• Bicistronic reporter assays to distinguish cap-dependent from IRES-dependent translation

• Electrophysiological studies (e.g., LTD measurements) to assess functional consequences of eIF4E modulation

• Context discrimination and other behavioral tests to examine cognitive effects

These complementary approaches allow for comprehensive analysis of eIF4E-dependent processes from molecular to behavioral levels.

How do insights from mouse eIF4E studies inform potential therapeutic strategies?

Mouse studies have revealed several promising therapeutic avenues:

• Cancer: Targeting eIF4E could impair cancer growth while minimally affecting normal physiological processes due to the excess eIF4E threshold

• Fragile X Syndrome: Inhibiting eIF4E-eIF4G interactions normalizes both synaptic plasticity and behavioral phenotypes

• Other neurological disorders: eIF4E modulation affects synaptic function and may be relevant for conditions with dysregulated protein synthesis

The differential requirements for eIF4E in normal physiology versus pathological states creates potential therapeutic windows for selective intervention.

What is the relationship between eIF4E and signaling pathways in mouse models?

eIF4E function is regulated by and influences several signaling pathways:

• mTORC1 pathway: Phosphorylates and inhibits 4E-BP2, releasing eIF4E so it can bind to eIF4G

• ERK pathway: Controls translation initiation via phosphorylation of eIF4E by MNK1 and MNK2 kinases

• Rac1-PAK-cofilin pathway: Connected to eIF4E via CYFIP1, regulating actin dynamics

• Metabotropic glutamate receptor pathway: Activates translation through eIF4E-dependent mechanisms

Understanding these regulatory relationships provides opportunities to indirectly modulate eIF4E function through established drug targets.