ECI1 Human

What is ECI1 and what is its primary function in human metabolism?

ECI1 (Enoyl-CoA Delta Isomerase 1) is a key mitochondrial enzyme involved in the beta-oxidation of unsaturated fatty acids. It belongs to the hydratase/isomerase superfamily and catalyzes the transformation of both 3-cis and 3-trans-enoyl-CoA esters into 2-trans-enoyl-CoA intermediates during the stepwise degradation of unsaturated fatty acids. This isomerization step is essential for the proper processing of these fatty acids through the beta-oxidation pathway. The enzyme plays a crucial role in energy metabolism, particularly when the body relies on fat metabolism during fasting or high-fat dietary conditions .

How is the ECI1 gene organized in the human genome?

The ECI1 gene is located on chromosome 16 in humans and encodes the mitochondrial enoyl-CoA isomerase protein. Several transcript variants have been described through alternative splicing mechanisms. The gene has been assigned various identifiers including HGNC: 2703, NCBI Gene: 1632, and Ensembl: ENSG00000167969. Previously, the gene was known by the symbol DCI (Dodecenoyl-CoA Delta Isomerase). The protein product is recognized in the UniProtKB/Swiss-Prot database with the identifier P42126 .

What metabolic pathways involve ECI1 activity?

ECI1 plays a critical role in the fatty acid metabolism pathway, particularly in the processing of unsaturated fatty acids. During beta-oxidation, when the pathway encounters a double bond at an odd-numbered carbon position, ECI1 catalyzes the isomerization of the resulting 3-cis or 3-trans-enoyl-CoA intermediates to the 2-trans form, which can then continue through the standard beta-oxidation cycle. This enzyme activity is essential for the complete oxidation of mono- and polyunsaturated fatty acids. Gene Ontology annotations indicate that ECI1 possesses delta(3)-delta(2)-enoyl-CoA isomerase activity and functions as an intramolecular oxidoreductase .

What are the known protein-protein interactions of ECI1?

ECI1 interacts with various proteins in the mitochondrial fatty acid oxidation pathway. Its functional relationship with ECI2 (a second enoyl-CoA isomerase) is particularly significant, as research indicates these enzymes demonstrate functional redundancy. This redundancy is evidenced by the ability of ECI2 to compensate for ECI1 deficiency in knockout models. The compensation mechanism explains the relatively mild phenotype observed in ECI1-deficient mice compared to other fatty acid oxidation enzyme deficiencies. AUH (AU RNA binding methylglutaconyl-CoA hydratase) is noted as an important paralog of the ECI1 gene, suggesting potential functional relationships in metabolic pathways .

What experimental models are most effective for studying ECI1 function?

The most informative model systems for studying ECI1 function include knockout mouse models and cell culture systems with gene knockdown capabilities. ECI1-deficient knockout mice have been successfully developed and characterized, providing valuable insights into the in vivo consequences of ECI1 deficiency. These models allow researchers to observe physiological responses to various metabolic challenges such as fasting or high-fat diets. For cellular studies, fibroblast models with ECI1 deficiency, with or without additional ECI2 knockdown, have proven effective for investigating the molecular consequences of enzyme deficiency and compensatory mechanisms. These complementary approaches allow researchers to connect molecular-level observations with physiological outcomes .

Table 1: Comparison of Experimental Models for ECI1 Research

How can researchers distinguish between ECI1 and ECI2 activity in experimental settings?

Distinguishing between ECI1 and ECI2 activities presents a significant challenge due to their functional redundancy. The most effective approach involves a combination of genetic manipulation and biochemical analysis. Sequential knockdown experiments, as demonstrated in the cited research, can help separate the contributions of each enzyme. First, researchers should establish ECI1-deficient models (either cells or organisms), then perform targeted knockdown of ECI2 to observe the additional effects. Substrate specificity analysis may also be informative, as the two enzymes might have different preferences or kinetics for specific fatty acid substrates despite their overlapping functions. Measuring acylcarnitine profiles before and after ECI2 knockdown in ECI1-deficient backgrounds provides quantitative evidence of their relative contributions to fatty acid metabolism .

What methodological approaches best identify potential ECI1 deficiency in human subjects?

Based on research findings from ECI1-deficient mouse models, several methodological approaches can help identify potential ECI1 deficiency in human subjects. The most promising diagnostic strategy involves metabolic profiling, particularly measuring acylcarnitine species in blood samples. Specifically, elevated levels of C12:1 acylcarnitine appear to be a sensitive biomarker for ECI1 deficiency, especially when assessed after metabolic stress such as fasting or after consumption of an unsaturated fat-rich diet. In the mouse model, C12:1 acylcarnitine levels were significantly elevated in knockout animals (WT 0.03 μM vs. KO 0.09 μM, P<0.01) during fasting, with even more pronounced differences observed after an olive oil-rich diet (WT 0.01 μM vs. KO 0.04 μM, P<0.01). Blood glucose monitoring may provide a secondary indicator, as ECI1-deficient mice showed a trend toward hypoglycemia during fasting (WT 4.58 mM vs. KO 3.87 mM, P=0.09) .

How should researchers design metabolic challenge studies to investigate ECI1 function?

Designing effective metabolic challenge studies for ECI1 investigation requires careful consideration of the physiological conditions that place demands on fatty acid oxidation pathways. Based on existing research protocols, effective approaches include:

• Fasting protocol: Implement a controlled food withdrawal period (typically 16-24 hours for mice, adjusted appropriately for human studies) to induce reliance on fatty acid oxidation.

• Dietary intervention: Administer a diet rich in unsaturated fatty acids (such as an olive oil-based diet) to specifically challenge the pathways requiring ECI1 activity.

• Comprehensive sampling: Collect blood samples for analysis of multiple parameters, including glucose, ketone bodies (particularly β-hydroxybutyrate), and a complete acylcarnitine profile.

• Tissue analysis: When possible, analyze tissue samples for enzyme activity and substrate accumulation to complement blood biomarkers.

• Time course measurements: Perform measurements at multiple time points to capture the dynamic nature of metabolic responses.

Research indicates that metabolic stress conditions significantly amplify the biochemical signatures of ECI1 deficiency, making them essential components of effective experimental designs .

What are the critical factors in interpreting acylcarnitine profiles in ECI1 research?

Interpreting acylcarnitine profiles in ECI1 research requires attention to several critical factors:

• Substrate specificity: C12:1 acylcarnitine appears to be the most sensitive biomarker for ECI1 deficiency, likely reflecting the accumulation of specific unsaturated fatty acid intermediates that require ECI1 for further processing.

• Contextual evaluation: Acylcarnitine levels should be evaluated in the context of the metabolic state (fed, fasted) and dietary composition (particularly unsaturated fat content).

• Comparative analysis: Results must be compared against appropriate controls matched for age, sex, and metabolic conditions.

• Functional redundancy consideration: The relatively modest elevations in acylcarnitines observed in ECI1 deficiency (compared to other fatty acid oxidation disorders) likely reflect the compensatory activity of ECI2.

• Pattern recognition: The pattern of multiple acylcarnitine species, rather than a single marker, may provide more comprehensive insights into pathway disruptions.

Researchers should note that the mild phenotypic presentation of ECI1 deficiency in mouse models suggests that human cases might also present with subtle biochemical abnormalities that could be missed without appropriate metabolic stress testing and careful analytical approaches .

How can researchers effectively control for ECI2 compensation in ECI1 studies?

Controlling for ECI2 compensation represents a significant methodological challenge in ECI1 research. Based on the reported research approaches, effective strategies include:

• Genetic manipulation: Implement simultaneous or sequential knockdown of both ECI1 and ECI2 to eliminate compensatory effects. This approach revealed that knockdown of ECI2 in ECI1-deficient fibroblasts caused a more pronounced accumulation of C12:1 acylcarnitine when incubated with unsaturated fatty acids (12-fold increase, P<0.05).

• Dose-dependent inhibition: Utilize graduated inhibition of ECI2 to establish a relationship between remaining enzyme activity and metabolic consequences.

• Tissue-specific analysis: Examine compensation effects across different tissues, as ECI2 expression and compensation capacity may vary.

• Substrate loading tests: Challenge the system with specific unsaturated fatty acid substrates that require isomerase activity to reveal the full metabolic impact when compensation is impaired.

• Temporal analysis: Investigate whether compensation mechanisms change over time, which might reveal progressive metabolic adaptations.

This multi-faceted approach allows researchers to distinguish primary effects of ECI1 deficiency from those masked by ECI2 compensation .

What is the predicted clinical presentation of human ECI1 deficiency based on animal models?

Based on animal model research, human ECI1 deficiency would likely present with a relatively mild clinical phenotype compared to other fatty acid oxidation disorders. The predicted presentation includes:

• Fasting hypoglycemia: Mild to moderate reductions in blood glucose levels during fasting or metabolic stress.

• Abnormal acylcarnitine profile: Characteristic elevations in unsaturated acylcarnitines, particularly C12:1 acylcarnitine, which becomes more pronounced during fasting or after unsaturated fat consumption.

• Relative absence of ketosis: Normal ketone body production (β-hydroxybutyrate levels) despite evidence of impaired fatty acid oxidation.

• Exacerbation with dietary challenges: Symptoms and biochemical abnormalities that become more evident with dietary unsaturated fat loading or during catabolic stress.

• Variable tissue involvement: Potential for tissue-specific manifestations depending on the relative importance of ECI1 versus ECI2 in different organs.

The mild phenotype observed in knockout mice suggests that human cases might be subclinical or present only during metabolic stress, which may explain why ECI1 deficiency has not yet been identified in humans despite the characterization of most other fatty acid oxidation disorders .

How does ECI1 research contribute to understanding broader fatty acid metabolism disorders?

ECI1 research provides valuable insights into the complexity of fatty acid metabolism disorders through several important contributions:

• Pathway redundancy: The functional overlap between ECI1 and ECI2 demonstrates how metabolic pathways incorporate redundancy to maintain essential functions, explaining the variable severity of different enzyme deficiencies.

• Biomarker discovery: The identification of specific acylcarnitine species associated with ECI1 deficiency helps refine diagnostic approaches for all fatty acid oxidation disorders.

• Stress-induced phenotypes: The research highlights how metabolic stress testing can reveal latent enzymatic deficiencies that remain compensated under normal conditions.

• Structure-function relationships: Understanding the specific catalytic role of ECI1 in processing unsaturated fatty acids illuminates the molecular basis of substrate specificity in beta-oxidation.

• Evolutionary conservation: The presence of multiple isomerases with overlapping functions reflects the evolutionary importance of maintaining unsaturated fatty acid metabolism.

These insights contribute to a more nuanced understanding of fatty acid metabolism disorders and may improve diagnostic and therapeutic approaches for the broader category of inborn errors of metabolism .

What genomic approaches might help identify human ECI1 variants of clinical significance?

Advancing genomic technologies offer promising approaches to identify human ECI1 variants of potential clinical significance:

• Targeted sequencing in at-risk populations: Screen individuals with unexplained hypoglycemia, mild fatty acid oxidation abnormalities, or characteristic acylcarnitine profiles for ECI1 variants.

• Whole exome/genome analysis: Include ECI1 in gene panels analyzing metabolic disorders, particularly in cases with partial or atypical presentations of fatty acid oxidation defects.

• Functional genomics: Develop high-throughput systems to characterize the functional impact of identified variants using cellular models and enzymatic assays.

• Population-based screening: Examine population databases for ECI1 variants with potential functional consequences, focusing on frequencies in different ethnic groups.

• Genotype-phenotype correlation studies: Connect specific genetic variants to biochemical profiles and clinical outcomes to establish pathogenicity and clinical relevance.

This multi-dimensional genomic approach offers the best opportunity to identify humans with ECI1 deficiency, which remains unrecognized despite the enzyme's important role in fatty acid metabolism .

What experimental designs would best elucidate the regulatory mechanisms of ECI1 expression?

Investigating the regulatory mechanisms controlling ECI1 expression requires sophisticated experimental designs that address multiple levels of gene regulation:

• Promoter analysis: Characterize the ECI1 promoter region to identify binding sites for transcription factors associated with metabolic regulation, particularly those responding to fatty acid levels and energy status.

• Epigenetic profiling: Map DNA methylation patterns and histone modifications across the ECI1 locus under different metabolic conditions to identify epigenetic regulatory mechanisms.

• Transcriptional regulation studies: Use reporter gene assays to identify regulatory elements and transcription factors that modulate ECI1 expression in response to metabolic signals.

• Post-transcriptional regulation: Investigate microRNA targeting, RNA stability determinants, and alternative splicing mechanisms that might influence ECI1 expression levels.

• Protein stability studies: Examine post-translational modifications and protein degradation pathways that regulate ECI1 protein levels and activity.

• Tissue-specific expression analysis: Compare regulatory mechanisms across different tissues to understand the basis for differential expression patterns.

These comprehensive approaches would provide insights into how ECI1 expression is modulated in different physiological states and how dysregulation might contribute to metabolic dysfunction .