NIP7 Human
Description
Introduction to NIP7 Human
NIP7 (Nucleolar Interacting Protein 7) is a conserved eukaryotic protein critical for ribosome biogenesis. In humans, it is encoded by the NIP7 gene located on chromosome 16 and functions in the maturation of ribosomal RNA (rRNA) and assembly of ribosomal subunits . NIP7 interacts with pre-ribosomal particles in the nucleolus and dissociates as subunits mature, playing a non-redundant role in 40S subunit biogenesis . Dysregulation of NIP7 disrupts rRNA processing, leading to imbalances in ribosomal subunit ratios (40S/60S) and impaired cell proliferation .
Key Functional Roles
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Pre-rRNA Processing: NIP7 depletion causes accumulation of 26S and 21S pre-rRNAs and reduced 34S pre-rRNA levels, indicating delayed cleavage at site 2 (critical for 18S rRNA maturation) .
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Ribosomal Subunit Biogenesis: NIP7 is required for 40S subunit production in humans, unlike its yeast ortholog, which primarily supports 60S subunit synthesis .
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RNA Binding: Recombinant NIP7 binds structured RNAs, particularly poly-AU sequences, suggesting a role in stabilizing rRNA intermediates .
Protein Interactions
Knockdown Studies
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Cell Proliferation: siRNA-mediated NIP7 downregulation in HEK293, MCF10A, and HeLa cells reduced proliferation rates by 40–60% .
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Cell Cycle Effects: Increased G0/G1 phase cells (e.g., 63% in HEK293 vs. 54% in controls) and reduced S-phase entry .
Pre-rRNA Processing Defects
Subcellular Localization
Comparative Analysis: Human vs. Yeast NIP7
Clinical and Evolutionary Implications
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Disease Links: NIP7 interacts with SBDS, mutations of which cause Shwachman-Diamond syndrome—a bone marrow failure disorder linked to ribosome dysfunction .
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Conservation: The C-terminal PUA domain (RNA-binding) is conserved across eukaryotes and archaea, underscoring its essential role in ribosome assembly .
Open Research Questions
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Mechanistic Role in Site 2 Cleavage: How NIP7 coordinates with endonucleases (e.g., U3 snoRNP) remains unclear .
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Therapeutic Potential: Could modulating NIP7 activity alleviate ribosomopathies or cancer (e.g., diseases with 40S subunit deficits)?
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RNA Specificity: Structural basis for poly-AU binding and its relevance to rRNA folding .
Product Specs
Q&A
What is the human NIP7 protein and what is its primary cellular function?
Human NIP7 is a conserved protein required for accurate pre-rRNA processing and ribosome biosynthesis. It is particularly involved in the maturation of 18S rRNA and 40S ribosomal subunit biogenesis. NIP7 is restricted to the nuclear compartment and co-sediments with complexes in the 40S-80S range, suggesting an association with nucleolar pre-ribosomal particles . Downregulation of NIP7 affects cell proliferation, consistent with its crucial role in rRNA biosynthesis in human cells .
What is the domain architecture of the human NIP7 protein?
The human NIP7 protein ranges from 160-180 amino acid residues and exhibits a two-domain architecture. The N-terminal domain consists of a five-stranded antiparallel β-sheet surrounded by three α-helices and a 3₁₀ helix. The C-terminal region contains the conserved PUA domain (named after Pseudo-Uridine synthases and Archaeosine-specific transglycosylases), which is a mixed β-sheet domain composed of strands β8 to β12, one α-helix, and a 3₁₀ helix . The PUA domain mediates NIP7's interaction with RNA .
How is NIP7 conserved across species?
NIP7 is highly conserved from archaea to humans, indicating its fundamental importance in cellular processes. Both the archaeal and eukaryotic NIP7 proteins can bind specifically to polyuridine RNA sequences, although through different structural mechanisms . In archaeal NIP7 (such as from Pyrococcus abyssi), positively charged residues (R151, R152, K155, and K158) in the PUA domain mediate RNA interaction. In contrast, eukaryotic NIP7 orthologues have these positions occupied by mostly hydrophobic residues (A/G160, I161, F164, and A167) .
How does NIP7 affect pre-rRNA processing in human cells?
Downregulation of NIP7 in human cells causes specific defects in pre-rRNA processing, leading to an imbalance in the 40S/60S ribosomal subunit ratio . Research has identified distinct molecular consequences:
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Decreased concentration of the 34S pre-rRNA
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Increased concentrations of the 26S and 21S pre-rRNAs
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Particularly slower processing at site 2 in NIP7-depleted cells
These findings demonstrate that NIP7 is specifically required for proper maturation of the 18S rRNA, which is essential for 40S ribosomal subunit formation .
What protein complexes does NIP7 associate with in human cells?
Human NIP7 participates in multiple protein-protein interactions essential for its function:
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Interacts with Nop132, the putative ortholog of S. cerevisiae Nop8p, which is also involved in ribosome biogenesis
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Found in complexes isolated by affinity-tagged purification of RPS19, a component of the 40S subunit essential for its synthesis
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Associates with parvulin (Par14), a peptidyl-prolyl cis-trans isomerase required for pre-rRNA processing
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Interacts with SBDS (Shwachman-Bodian-Diamond syndrome) protein in the yeast two-hybrid system, with GST-SBDS being able to pull down NIP7 from HEK293 cell extracts
What are effective methods for downregulating NIP7 in experimental settings?
Researchers can employ several approaches to downregulate NIP7 expression:
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Stable transfection using shRNA:
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Transient transfection using siRNA:
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For HeLa and MCF10A cells:
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For HEK293 cells:
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Control conditions:
How can researchers analyze pre-rRNA processing defects in NIP7-depleted cells?
The following methodological approaches are effective for analyzing pre-rRNA processing defects:
When analyzing data, researchers should pay particular attention to:
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Changes in the relative abundance of specific pre-rRNA intermediates
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Alterations in the 40S/60S subunit ratio
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Impact on cell proliferation rates, which typically decrease with NIP7 depletion
How does the PUA domain of NIP7 mediate RNA interaction?
The PUA domain in the C-terminal region of NIP7 is crucial for RNA binding. Structural alignment of the P. abyssi NIP7 (PaNip7) PUA domain with the RNA-interacting surface of archaeosine tRNA-guanine transglycosylase (ArcTGT) PUA domain has revealed important insights :
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In archaeal PUA domains, positively charged residues (R151, R152, K155, and K158) facilitate RNA interaction
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In eukaryotic NIP7 orthologues, these positions are occupied by mostly hydrophobic residues (A/G160, I161, F164, and A167)
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Despite these structural differences, both archaeal and eukaryotic NIP7 proteins bind specifically to polyuridine sequences
Site-directed mutagenesis experiments have confirmed that specific residues within the PUA domain are required for RNA interaction .
What is the significance of NIP7's RNA binding specificity?
NIP7's specific binding to polyuridine RNA sequences has important functional implications:
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This specificity is conserved between archaeal and eukaryotic NIP7 proteins, suggesting evolutionary importance
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The preference for polyuridine may direct NIP7 to specific regions of pre-rRNAs during processing
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This RNA binding property is likely essential for NIP7's role in pre-rRNA processing and 40S ribosomal subunit assembly
What is the relationship between NIP7 and genetic disorders?
While direct causative relationships between NIP7 mutations and specific genetic disorders haven't been firmly established in the provided research data, important associations exist:
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NIP7 interacts with the SBDS protein, mutations in which cause Shwachman-Bodian-Diamond syndrome
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Loss-of-function mutations in trans-acting factors involved in ribosome biogenesis can lead to genetic syndromes
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Given NIP7's essential role in ribosome biogenesis, particularly in 18S rRNA maturation, its dysfunction could potentially contribute to ribosomopathies (disorders of ribosome formation or function)
How does NIP7 deficiency impact cellular physiology?
Downregulation of NIP7 affects cellular function through several mechanisms:
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Impaired pre-rRNA processing:
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Reduced cell proliferation:
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Potential impact on gene expression:
What are promising future research directions for understanding NIP7 function?
Several promising research avenues could further elucidate NIP7's function:
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Structural studies:
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Determine the crystal structure of human NIP7 bound to RNA
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Compare with the known archaeal structures to understand mechanistic differences
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Interactome mapping:
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Comprehensive identification of NIP7-interacting proteins
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Temporal analysis of interaction dynamics during ribosome biogenesis
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Disease-association studies:
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Investigation of potential NIP7 mutations in patients with undiagnosed ribosomopathies
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Development of animal models with conditional NIP7 depletion
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Therapeutic targeting:
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Exploration of NIP7 as a potential target in diseases characterized by dysregulated ribosome biogenesis, such as certain cancers
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What methodological challenges exist in studying human NIP7?
Researchers face several challenges when studying NIP7:
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Data interpretation complexities:
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Pre-rRNA processing involves multiple parallel pathways
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Need for streamlined data analysis, as shown in tables like this example from related research:
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| ID | Group | I01 | I02 | I03 | I04 | I05 | I06 | Sum |
|---|---|---|---|---|---|---|---|---|
| s01 | A | 3 | 4 | 5 | 1 | 2 | 3 | 18 |
| s02 | A | 2 | 5 | 3 | 2 | 1 | 1 | 14 |
| s03 | A | 4 | 5 | 4 | 3 | 1 | 1 | 18 |
| s04 | B | 2 | 1 | 5 | 4 | 3 | 3 | 18 |
| s05 | B | 2 | 1 | 2 | 5 | 2 | 1 | 13 |
| s06 | B | 2 | 3 | 3 | 4 | 2 | 1 | 15 |
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Technical limitations:
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Ensuring complete NIP7 knockdown without off-target effects
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Distinguishing direct from indirect effects on pre-rRNA processing
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Temporal resolution of pre-rRNA processing events
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Product Science Overview
Nuclear Import 7 Homolog, also known as NIP7, is a protein that plays a crucial role in the biogenesis of the 60S ribosomal subunit. This protein is essential for the proper assembly and function of ribosomes, which are the cellular machinery responsible for protein synthesis. The recombinant form of this protein, produced in Escherichia coli, is often used in research to study its function and interactions.
The recombinant Human Nuclear Import 7 Homolog is a single, non-glycosylated polypeptide chain consisting of 188 amino acids. It has a molecular mass of approximately 21.5 kDa . The protein solution typically contains 20mM Tris-HCl buffer (pH 8.0), 20% glycerol, and 0.1M NaCl . For long-term storage, it is recommended to add a carrier protein such as 0.1% Human Serum Albumin (HSA) or Bovine Serum Albumin (BSA) to prevent degradation .
NIP7 is involved in the nuclear import of ribosomal proteins and other macromolecules. It interacts with import receptors and nucleoporins to facilitate the transport of these molecules through the nuclear pore complex (NPC). This process is vital for maintaining cellular function and ensuring the proper assembly of ribosomes .
The nuclear import of proteins like NIP7 involves several steps:
- Recognition and Binding: NIP7 is recognized by import receptors such as importin 7. These receptors bind to the nuclear localization signals (NLS) present on the NIP7 protein .
- Transport through NPC: The import receptor-NIP7 complex is transported through the NPC. Nucleoporins, such as Nup358, play a crucial role in this process by interacting with the import receptor and facilitating the translocation of the complex into the nucleus .
- Release and Recycling: Once inside the nucleus, the NIP7 protein is released from the import receptor, which is then recycled back to the cytoplasm to transport more proteins .
Recombinant NIP7 is widely used in research to study its role in ribosome biogenesis and nuclear import. It is also used to investigate the interactions between import receptors and nucleoporins, as well as the mechanisms underlying nuclear transport. Understanding these processes can provide insights into various cellular functions and diseases related to ribosome assembly and protein synthesis.
