NAA30 Human

What is human NAA30 and what is its primary function?

Human NAA30 is the catalytic subunit of the NatC complex, a major N-terminal acetyltransferase (NAT) enzyme that catalyzes the irreversible co-translational N-terminal acetylation of proteins. This protein modification is defined by the addition of an acetyl group to the N-terminus of a protein. The NatC complex is estimated to N-terminally acetylate approximately 20% of the human proteome . The acetylation transforms the positively charged N-terminus into a hydrophobic handle, which can significantly impact protein function, stability, and localization . NAA30 contains the recognizable Gcn5-related N-acetyltransferase (GNAT) fold that is characteristic of acetyltransferases and harbors the enzymatic activity of the NatC complex .

How is the NatC complex structured in humans?

The human NatC complex is a heterotrimeric complex consisting of three subunits:

• NAA30 (previously known as MAK3 in yeast): The catalytic subunit that possesses the acetyltransferase activity

• NAA35 (MAK10 in yeast): Functions as the ribosomal anchor

• NAA38 (MAK31 in yeast): Serves as an auxiliary subunit that enhances complex function

This tripartite structure is conserved from yeast to humans, although human NAA30 has a N-terminal extension that is not present in the yeast homolog or other catalytic human NAT subunits . Recent research indicates that NAA38 broadens NatC's substrate specificity and increases its thermostability by repositioning NAA30 . A crystal structure of the yeast NatC complex has been determined, revealing the molecular interactions between the three subunits, though a detailed structure of the human NatC complex has not yet been reported .

What is the substrate specificity profile of human NAA30?

Human NatC, with NAA30 as its catalytic subunit, has a distinct substrate specificity profile:

Human NatC co-translationally acetylates the N-termini of proteins that begin with methionine followed by a hydrophobic or amphipathic residue (Met-Leu/Ile/Phe/Trp/Val/Met/Lys) . The auxiliary subunit NAA38 has been shown to broaden this substrate specificity . This substrate profile is similar to that observed in yeast NatC, highlighting the evolutionary conservation of this enzymatic function .

What cellular processes are dependent on NAA30-mediated N-terminal acetylation?

NAA30-mediated N-terminal acetylation has been implicated in several critical cellular processes:

• Mitochondrial function: Depletion of NAA30 disrupts mitochondrial function in human cancer cells and reduces levels of mitochondrial matrix proteins .

• Protein targeting: N-terminal acetylation by NatC is required for protein complex formation (Ubc12-Dcn1) and subcellular targeting to:

  • The Golgi apparatus (Arl3 and Grh1)

  • The inner nuclear membrane (Trm1-II)

• Protein stability: NatC plays a role in shielding the proteome from degradation. Loss of NAA30-mediated N-terminal acetylation can lead to increased protein degradation through the ubiquitin-proteasome system .

• Energy metabolism: In various organisms, NatC is involved in energy regulation and normal growth patterns .

What experimental models are available to study human NAA30 function?

Several experimental models have been developed to study human NAA30 function:

• Yeast models: The study by Drazic and Varland demonstrates that human NAA30 can functionally replace yeast MAK3/NAA30 in rescue experiments. This provides a powerful system to study human NAA30 variants using growth assays under various stress conditions . The authors showed that yeast cells lacking MAK3/NAA30 grow poorly in non-fermentable carbon sources and other stress conditions, and that human NAA30 can rescue these growth defects .

• Drosophila melanogaster models: Studies in fruit flies have shown that NatC is essential for longevity, fertility, and prevention of age-dependent motility loss. The decreased longevity and motility in NatC deletion flies were rescued by muscle-specific overexpression of UbcE2M (the fruit fly homologue of human NEDD8-conjugating enzymes), supporting a role for NatC in protecting against protein degradation and in normal muscle development .

• Caenorhabditis elegans models: In C. elegans, NatC regulates the balance between reproductive growth and stress tolerance in response to nutrients and stressors .

• Human cell lines: Depletion of NAA30 in human cancer cells disrupts mitochondrial function and morphology, and increases lysosomal content and cell granularity .

• Arabidopsis thaliana models: In this plant model, NatC (specifically NAA30) is required for efficient photosynthesis and growth .

How can yeast-based functional assays be used to study human NAA30 variants?

Yeast-based functional assays represent a powerful approach to study potential pathogenic variants of human NAA30:

• Complementation studies: Despite limited sequence identity (19.8%) between yeast Mak3 and human NAA30, human NAA30 can functionally replace yeast MAK3/NAA30. This functional conservation enables the use of yeast as a model to study human NAA30 variants .

• Growth assays in stress conditions: The researchers demonstrated that liquid growth assays are more sensitive than plate-based methods for detecting subtle growth defects in yeast. This methodology can be applied in a high-throughput manner to test different stress conditions and evaluate the functional impact of human NAA30 variants .

• Strain dependency: The rescue ability of human NAA30 depends on the genetic background of the yeast strain, highlighting the importance of choosing appropriate yeast strains for these studies .

• Biochemical validation: Immunoblotting can be used to confirm stable expression of human NAA30 in yeast, ensuring that any observed phenotypes are due to functional differences rather than expression issues .

This yeast system provides a simple and efficient approach to rapidly assess the functionality of human NAA30 variants in vivo before pursuing more complex and time-consuming mammalian models .

What are the implications of NAA30 variants in human diseases?

While extensive research has been conducted on the pathogenic variants of other NAT genes (like NAA10 and NAA15), information on NAA30 variants in human diseases is emerging:

• Neurodevelopmental disorders: A heterozygous nonsense variant in the NAA30 gene was identified in an individual presenting with global developmental delay, autism spectrum disorder, and a tracheal cleft . This suggests that NAA30 dysfunction may contribute to neurodevelopmental conditions.

• Potential mechanism: The clinical manifestations associated with NAA30 variants might be linked to aberrant N-terminal acetylation of critical substrates involved in neurodevelopment and other physiological processes .

• Comparison to other NAT-related disorders: Other NAT genes like NAA10, NAA15, and NAA20 have established links to human disorders:

  • NAA10 variants cause Ogden syndrome and other NAA10-related syndromes characterized by developmental delay, intellectual disability, and cardiac anomalies .

  • NAA15 variants are associated with congenital heart disease .

  • NAA20 variants cause NAA20-related syndrome with intellectual disability, developmental delay, and microcephaly .

Given these precedents, it is likely that further research will uncover additional clinical implications of NAA30 variants. The yeast-based functional assay described by Drazic and Varland provides a valuable tool for evaluating the pathogenicity of newly identified NAA30 variants .

How does the NatC complex regulate mitochondrial function in human cells?

The regulation of mitochondrial function by the NatC complex involves several mechanisms:

• N-terminal acetylation of mitochondrial proteins: NatC acetylates the N-termini of certain mitochondrial matrix proteins, which affects their stability and function .

• Impact on mitochondrial morphology: Depletion of NAA30 in human cells disrupts mitochondrial morphology, suggesting a role in maintaining mitochondrial structure .

• Organelle-specific effects: The impact of NatC on mitochondrial function appears to be conserved across species. In yeast, all three NatC subunits are necessary for NatC-mediated N-terminal acetylation and proper mitochondrial function . Similarly, in human cells, NAA30 depletion results in reduced levels of mitochondrial matrix proteins .

• Protection from degradation: The NatC complex shields certain proteins from degradation, which may include key mitochondrial components. Loss of this protection in NAA30-deficient cells could contribute to mitochondrial dysfunction .

• Energy metabolism: Yeast cells lacking MAK3/NAA30 grow poorly in non-fermentable carbon sources, which require mitochondrial respiration for utilization, indicating that NAA30 is important for mitochondrial energy production .

What methodological considerations are important when studying NAA30 function across different experimental systems?

When studying NAA30 function across different experimental systems, researchers should consider:

What are the most pressing research questions regarding human NAA30 that remain to be addressed?

Several important questions about human NAA30 remain to be addressed:

• Structural basis of human NatC function: While the crystal structure of yeast NatC has been determined, a detailed structure of the human NatC complex has not yet been reported . Such structural information would provide important insights into how human NatC recognizes its substrates.

• Function of the N-terminal extension: Human NAA30 has a N-terminal extension that is neither present in other catalytic human NAT subunits nor in the yeast homolog Mak3. The function of this N-terminal segment remains unknown .

• Complete substrate specificity profile: Further research is needed to comprehensively identify all physiological substrates of human NAA30 and understand how substrate recognition contributes to various cellular processes.

• Tissue-specific roles: The tissue-specific functions of NAA30, particularly in the context of human development and disease, remain to be fully elucidated.

• Therapeutic potential: Whether modulation of NAA30 activity could have therapeutic applications for mitochondrial disorders or other conditions linked to dysregulated N-terminal acetylation requires investigation.

How might emerging technologies advance research on NAA30 and N-terminal acetylation?

Emerging technologies could significantly advance our understanding of NAA30:

• CRISPR-Cas9 genome editing: Generation of precise NAA30 variants in human cells and model organisms to study their functional consequences.

• Single-cell proteomics: Analysis of N-terminal acetylation patterns at the single-cell level to understand cell-to-cell variability and tissue-specific functions.

• Cryo-electron microscopy: Determination of high-resolution structures of the human NatC complex, potentially in complex with ribosomal components or substrates.

• Induced pluripotent stem cells (iPSCs): Generation of patient-derived iPSCs harboring NAA30 variants to study their impact on different cell types and developmental processes.

• High-throughput screening: Development of small molecule modulators of NAA30 activity for research tools and potential therapeutic applications.

• Computational modeling: Prediction of the functional impact of NAA30 variants and identification of novel substrates through machine learning approaches.