Critical and Differential Roles of eIF4A1 and eIF4A2 in B-Cell Development and Function

Academic Background

In mammalian cells, translation initiation is a crucial step in protein synthesis, where the eukaryotic initiation factor 4A (eIF4A) plays a pivotal role. eIF4A is an ATP-dependent RNA helicase that unwinds secondary structures in the 5’ untranslated region (5’ UTR) of mRNAs, enabling the 43S preinitiation complex (PIC) to scan the 5’ UTR to locate the start codon. eIF4A has two highly homologous isoforms: eIF4A1 and eIF4A2. Although they can interchangeably participate in the formation of the eIF4F complex in vitro, their biological functions and molecular mechanisms in vivo remain incompletely understood.

B cells are essential components of the immune system, responsible for producing antibodies to combat infections and diseases. The development and function of B cells are governed by intricate post-transcriptional regulatory mechanisms, including RNA-binding proteins (RBPs), microRNAs (miRNAs), alternative splicing, and others. The specific functions of eIF4A1 and eIF4A2 in B cells remain unclear, particularly their roles in B-cell development, activation, and antibody production. Furthermore, eIF4A1 is significantly upregulated in B-cell lymphoma, suggesting a potential role in B-cell-related diseases.

Paper Source

This study was conducted by Ying Du, Jun Xie, Dewang Liu, and colleagues from Xiamen University School of Life Sciences, The Scripps Research Institute, and other institutions. The paper was published online in the journal Cellular & Molecular Immunology on November 8, 2024, with the DOI 10.1038/s41423-024-01234-x.

Research Process and Results

1. Critical Roles of eIF4A1 and eIF4A2 in B-Cell Development

The authors first analyzed the expression patterns of eIF4A1 and eIF4A2 during B-cell development and activation using RT-qPCR and immunoblotting. eIF4A1 was highly expressed in the early stages of B-cell development (e.g., Hardy Fraction C) but had low expression in mature B cells. In contrast, eIF4A2 expression gradually increased throughout development, peaking in mature recirculating B cells.

To further elucidate the functions of eIF4A1 and eIF4A2, the team generated conditional knockout mice (eIF4A1fl/fl;Mb1Cre and eIF4A2fl/fl;Mb1Cre). eIF4A1 deficiency resulted in a developmental block at Hardy Fractions B and C; on the other hand, eIF4A2 deficiency caused a block at Fraction D. Therefore, eIF4A1 and eIF4A2 play essential and distinct roles in different stages of early B-cell development.

2. Roles of eIF4A1 and eIF4A2 in T-Cell-Dependent Antibody Responses

To investigate their roles in immune responses, the team used CD19Cre mice to bypass early developmental defects. Both eIF4A1 and eIF4A2 deficiencies significantly reduced germinal center B-cell (GCB) and plasma cell numbers while impairing antigen-specific antibody production. These results demonstrate that both eIF4A1 and eIF4A2 are indispensable for T-cell-dependent antibody responses.

3. Differential Roles in T-Cell-Independent Antibody Responses

The roles of these factors were further examined in T-cell-independent (TI) antibody responses. eIF4A1 was essential for TI-1 antibody responses (e.g., NP-LPS-induced responses), while eIF4A2 was required for both TI-1 and TI-2 responses (e.g., NP-Ficoll-induced responses). This indicates that eIF4A1 and eIF4A2 have distinct roles in different types of TI responses.

4. Regulation of B-Cell Proliferation by eIF4A1 and eIF4A2

The knockout of either eIF4A1 or eIF4A2 did not affect B-cell activation markers but significantly impaired cell proliferation. Cell cycle analysis revealed that both were critical for the G1 to S phase transition, highlighting their importance in promoting B-cell proliferation.

5. eIF4A2 Regulates Biogenesis of the 40S Ribosomal Subunit

Using polysome profiling, the researchers found that eIF4A1 deficiency reduced the global translation rate, whereas eIF4A2 deficiency specifically diminished 40S ribosomal subunit levels. eIF4A2 was shown to regulate the maturation of 18S rRNA, a critical component of the 40S ribosome. Its deficiency impaired 18S rRNA processing, leading to unassembled ribosomal proteins (RPS) being degraded.

6. eIF4A1 Regulates Cell Cycle Progression via GINS4 and Other Targets

RNA-seq and mass spectrometry analyses revealed that eIF4A1 deficiency downregulated the translation of several cell cycle genes, including GINS4, a key component of the replication complex. eIF4A1 promotes GINS4 translation by resolving complex secondary structures in its 5’ UTR. CRISPR/Cas9 knockout experiments further suggested that GINS4 is a major mediator of eIF4A1’s role in cell cycle regulation.

Conclusions and Significance

This study uncovers the critical and distinct roles of eIF4A1 and eIF4A2 in B-cell development and function. eIF4A1 primarily regulates global protein synthesis and the cell cycle, while eIF4A2 controls 40S ribosome biogenesis via 18S rRNA maturation. These findings not only deepen our understanding of the molecular functions of eIF4A isoforms but also provide potential therapeutic targets for B-cell-related diseases.

Research Highlights

1. Key Findings: eIF4A1 and eIF4A2 exhibit distinct functions in B cells, with eIF4A2’s role in 40S ribosome biogenesis being particularly novel.
2. Innovative Methods: The use of conditional knockout mouse models and techniques such as polysome profiling offers comprehensive insights into eIF4A isoform functions.
3. Clinical Relevance: The regulatory mechanisms of eIF4A1 and eIF4A2 may offer new therapeutic avenues for B-cell-related diseases such as lymphoma.

Additional Insights

The study provides detailed experimental protocols and data analysis workflows, such as RNA-seq, mass spectrometry, and polysome profiling, serving as valuable resources for researchers in related fields. The RNA-seq datasets generated in this study are also publicly available for further analysis.

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This research not only advances our understanding of eIF4A1 and eIF4A2’s biological roles but also lays the groundwork for future studies in immunology and cancer biology.