RPS4X Human
Description
Introduction to RPS4X Human
RPS4X (Ribosomal Protein S4 X-linked) is a gene located on the human X chromosome (Xq13.2) that encodes ribosomal protein S4, a component of the 40S ribosomal subunit critical for protein synthesis. Unlike most X-linked genes, RPS4X escapes X-inactivation, allowing females to express two functional copies . Its homolog, RPS4Y, resides on the Y chromosome and produces a distinct isoform, though both proteins are functionally equivalent . RPS4X is implicated in ribosomal assembly, translation regulation, and has emerged as a biomarker in various cancers .
Gene Structure and Evolution
RPS4X spans 7 exons with conserved intron positions across species. The 5' flanking regions of RPS4X and the mouse ortholog Rps4 show high similarity, while RPS4Y diverges upstream of the transcription start site . This suggests evolutionary conservation of regulatory elements for RPS4X.
| Feature | RPS4X | RPS4Y |
|---|---|---|
| Chromosomal Location | Xq13.2 | Yp11.3 |
| Exons | 7 | 7 |
| X-Inactivation | Escapes | N/A (Y-linked) |
| Amino Acid Differences | 19 from RPS4Y (out of 263) | 19 from RPS4X |
| Tissue Expression | Ubiquitous | Ubiquitous (lower abundance) |
Key Insight: RPS4X and RPS4Y diverged before placental mammal radiation, with RPS4Y absent in many species . Chicken S4, encoded by a single gene, exhibits recombination-like features between X/Y homologs .
Functional Role in Ribosome Biogenesis
RPS4X is essential for ribosome assembly and translation. The 40S subunit, containing RPS4X, binds mRNA and tRNA during protein synthesis . Studies show:
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Interchangeability: RPS4X and RPS4Y rescue a temperature-sensitive ribosome defect in hamster cells, confirming functional equivalence .
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Translation Regulation: RPS4X interacts with Y-box binding protein-1 (YB-1) to modulate cisplatin resistance in cancer cells .
Controversies in Turner Syndrome Pathogenesis
Haploinsufficiency of RPS4X was hypothesized to contribute to Turner syndrome (45,X), but evidence remains inconclusive:
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Pro-Haploinsufficiency:
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Counterarguments:
RPS4X in Cancer: Diagnostic and Prognostic Biomarker
RPS4X overexpression correlates with poor outcomes in multiple cancers:
Key Finding: In ICC, high RPS4X expression predicts shorter survival (HR = 1.82, p < 0.001) .
Protein Interactions and Functional Partners
RPS4X interacts with ribosomal proteins and regulators, as mapped by STRING and co-expression networks:
| Interactor | Confidence Score | Functional Role |
|---|---|---|
| RPL18A | 0.999 | 60S subunit assembly |
| RPL19 | 0.999 | Ribosome maturation |
| RPS12 | 0.999 | 40S subunit stability |
| YB-1 | 0.85 | Translation regulation, cisplatin resistance |
Clinical and Research Implications
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Therapeutic Target: RPS4X/YB-1 complexes may be exploited to overcome cisplatin resistance in breast/ovarian cancers .
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Diagnostic Potential: High specificity for ICC detection using immunohistochemistry .
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Evolutionary Insights: Conservation across species highlights its critical role in translation machinery .
Product Specs
Q&A
What is the basic structure and function of human RPS4X?
Human RPS4X is an X-linked gene encoding the X isoform of ribosomal protein S4, a component of the 40S ribosomal subunit. The protein consists of 263 amino acids and plays an essential role in ribosome assembly and function . RPS4X is present in translationally active ribosomes and provides a fundamental function in protein synthesis. Unlike most X-linked genes, RPS4X escapes X-inactivation, resulting in expression from both X chromosomes in females . The gene is organized into seven exons, with intron positions that are conserved between humans and mice .
How do RPS4X and RPS4Y proteins differ structurally?
The human RPS4X and RPS4Y proteins, while functionally similar, differ at 19 of their 263 amino acid positions (approximately 7.2% difference) . Despite these differences, functional studies have demonstrated that both proteins are interchangeable in their essential roles . The genes encoding these proteins share high sequence similarity (82% identical at the DNA level), but diverge notably in their 5' flanking sequences, particularly upstream of the transcription start site . This structural divergence may contribute to differences in expression regulation between the two genes.
What is the chromosomal location and genomic context of RPS4X?
RPS4X maps to the long arm of the X chromosome (Xq) near the X-inactivation center but critically escapes X inactivation . Specifically, it is located in the vicinity of the phosphorylase kinase alpha 1 (PHKA1) gene and the X-inactive specific transcript (XIST) gene . The gene order has been determined as cen--RPS4X--PHKA1--XIST--DXS128E--tel, with the transcriptional orientation of RPS4X being cen--5-prime--3-prime--qter . This genomic context is particularly important for understanding its role in X-chromosome biology.
What techniques are most effective for measuring RPS4X expression levels?
For quantitative analysis of RPS4X expression, researchers commonly employ:
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Quantitative PCR (qPCR): As demonstrated in experimental studies, RPS4X mRNA expression can be measured using SYBR Green-based qPCR systems . Primer design is critical for specificity given the similarity to RPS4Y.
Recommended primer sequences:
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Forward: 5'-CCTGGATCTTTTGACGTGGT-3'
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Reverse: 5'-TTTCCTCGGGGAAGAGAAAT-3'
Reference gene (B2M) primers for normalization:
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Forward: 5'-TGACTTTGTCACAGCCCAAG-3'
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Reverse: 5'-AGCAAGCAAGCAGAATTTGG-3'
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Immunohistochemistry (IHC): For protein-level detection in tissue samples, IHC using isoform-specific antibodies has proven effective, with Integrated Optical Density (IOD) values providing quantitative measurements .
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RNA interference: For functional studies, endoribonuclease-prepared siRNA (esiRNA) has demonstrated high specificity in targeting RPS4X without affecting RPS4Y expression despite their sequence similarity .
How can researchers effectively distinguish between RPS4X and RPS4Y in experimental settings?
Discriminating between these similar isoforms requires:
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Isoform-specific antibodies: The development of antisera specific to either S4X or S4Y allows for differential detection of these proteins in cellular samples . These antibodies can identify the relative abundance of each isoform in ribosomes from various tissue types.
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Sequence-specific oligonucleotides: Designing primers or probes that target the 18% sequence divergence between the genes ensures specificity in nucleic acid-based detection methods .
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Expression pattern analysis: In cells with varying sex chromosome constitutions (46,XY; 49,XYYYY; 49,XXXXY), the relative abundance of S4Y to S4X changes proportionally to the number of Y chromosomes present, providing a method for validation of isoform-specific detection .
What is the evidence for RPS4X involvement in Turner syndrome?
Turner syndrome, characterized by monosomy X (45,X), has been hypothesized to involve RPS4X insufficiency:
How does RPS4X expression correlate with cancer prognosis?
RPS4X has demonstrated significant value as a cancer biomarker:
What are the current hypotheses explaining how RPS4X escapes X-inactivation?
RPS4X is among a minority of X-linked genes that escape X-inactivation. Several mechanisms have been proposed:
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Genomic context: The proximity of RPS4X to the X-inactivation center may create a chromosomal environment that protects it from the silencing mechanisms .
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Regulatory elements: The divergence in 5' flanking sequences between RPS4X and RPS4Y suggests unique regulatory elements that may contribute to the escape from X-inactivation .
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Evolutionary pressure: The essential nature of ribosomal proteins may have selected for mechanisms that ensure adequate expression levels in females, leading to evolutionary conservation of X-inactivation escape .
Unlike most X-linked genes, RPS4X is not dosage compensated, resulting in a unique expression pattern in male versus female cells . This escape from X-inactivation has significant implications for understanding sex-based differences in ribosome composition and function.
How does the ribosomal composition differ between males and females due to RPS4X/Y expression?
The differential expression of RPS4X and RPS4Y creates a sex-specific pattern in ribosomal composition:
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In male cells (46,XY), S4Y constitutes approximately 10-15% of the total S4 protein in ribosomes, with S4X making up the remainder . This creates a distinct male-specific ribosomal composition.
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In female cells, which lack RPS4Y but express RPS4X from both X chromosomes due to escape from X-inactivation, all S4 protein is of the X isoform .
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In cells with atypical sex chromosome constitutions, the ratio changes proportionally:
This sex-specific difference in ribosomal composition may have functional implications that remain to be fully elucidated but suggests that the ribosomes of human males and females are structurally distinct .
What evolutionary insights can be gained from studying RPS4X across species?
Comparative studies of RPS4 across species have revealed important evolutionary patterns:
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In mice, there is only an X-linked gene (Rps4) with no Y homolog, and the mouse S4 protein is identical to human S4X . Unlike human RPS4X, mouse Rps4 undergoes normal X-inactivation .
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In chickens, S4 is encoded by a single gene that is not sex-linked . Interestingly, the chicken S4 protein differs from human S4X by only four amino acid substitutions, all within a region encoded by a single exon. Three of these four substitutions are also present in human S4Y, suggesting potential recombination events in evolution .
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The conservation of intron positions across human RPS4X, RPS4Y, and mouse Rps4 genes indicates a shared ancestral origin despite their subsequent divergent evolution .
These evolutionary comparisons provide a framework for understanding how sex-specific ribosomal proteins may have evolved and suggest mechanisms for the origination of Y-linked genes from X-linked progenitors.
Product Science Overview
Ribosomal Protein 4X (RPS4X) is a component of the 40S subunit of cytoplasmic ribosomes, which are essential organelles responsible for protein synthesis in cells. Ribosomes consist of a small 40S subunit and a large 60S subunit, together composed of four RNA species and approximately 80 structurally distinct proteins . RPS4X belongs to the S4E family of ribosomal proteins and is unique in that it is encoded by more than one gene, specifically the X-linked RPS4X and the Y-linked RPS4Y .
RPS4X is a highly conserved protein that plays a crucial role in the assembly and function of the ribosome. It is involved in the translation of messenger RNA (mRNA) into proteins, a fundamental process for cellular life . The protein is composed of 263 amino acids and has a molecular weight of approximately 32 kDa . The structure of RPS4X includes an N-terminal His-tag, which facilitates its purification and study in recombinant form .
The expression of ribosomal proteins, including RPS4X, is tightly regulated to maintain the appropriate stoichiometry of ribosomal components. Feedback mechanisms exist to control the production of ribosomal proteins, ensuring that their levels are balanced with other ribosomal components . In humans, the expression of RPS4X is not subject to X-inactivation, meaning that both the X-linked and Y-linked versions of the protein are expressed .
Recombinant RPS4X is typically produced using bacterial expression systems, such as Escherichia coli (E. coli). The gene encoding RPS4X is cloned into an expression vector, which is then introduced into E. coli cells. The bacteria are cultured, and the recombinant protein is expressed and purified using chromatography techniques . The purified protein is often stored in a buffer containing Tris-HCl, glycerol, NaCl, and dithiothreitol (DTT) to maintain its stability .
RPS4X, like other ribosomal proteins, can undergo various post-translational modifications that may affect its function and interactions. These modifications include phosphorylation, methylation, and acetylation. Analytical techniques such as mass spectrometry, X-ray crystallography, and nuclear magnetic resonance (NMR) spectroscopy are used to study the structure and function of RPS4X .
RPS4X has been implicated in various cellular processes and diseases. For example, haploinsufficiency of the RPS4X gene has been suggested to play a role in Turner syndrome, although this hypothesis remains controversial . Additionally, dysregulated expression of ribosomal proteins, including RPS4X, has been observed in certain cancers and is predictive of disease progression .
