New insights into macular carotenoid accumulation and transport

Reviewed by Dr Johannes von Lintig.

Carotenoids are pigments that occur naturally and, as well as providing the red-to-orange colours of many fruits and vegetables, have important functions in the eye. One such carotenoid, beta-carotene, is an essential nutrient and the major precursor of vitamin A.

Beta-carotene is converted in the human body to two metabolites: 11-cis-retinal and all-transretinoic acid. Through those molecules, it affects major physiological functions (vision, cellular differentiation, metabolic control, reproduction and embryonic development and organogenesis), according to Dr Johannes von Lintig, a professorin the Department of Pharmacology at Case Western Reserve University, Cleveland, Ohio, United States.

In addition to (pro) vitamin A, other carotenoids are absorbed and accumulated in the central retina (the macula lutea) of the human eyes, namely lutein, zeaxanthin and meso-zeaxanthin. All of these substances reduce chromatic aberrations, increase visual acuity and prevent oxidative stress to photoreceptors.

The most important research finding regarding carotenoid and vitamin A metabolism is the association with numerous diseases, such as xerophthalmia; age-related macular degeneration; micro-ophthalmic syndromes; retinitis pigmentosa; and iris, retinal and choroidal coloboma.

An unanswered question

Although a greatdeal is known about vitamin A metabolism in the retina, Dr von Lintigwas interested in the mechanism of carotenoid accumulation in the macula. His laboratory identified a carotenoid scavenger gene,beta-carotene-oxygenase 2 (BCO2), which encodes a mitochondrial enzyme expressed in the inner membrane of the mitochondria that breaks down carotenoids, such as zeaxanthin, into smaller entities.

The enzyme is important in producing the colours seen in nature. As Dr von Lintig and colleagues from Columbia University, New York, US,recently demonstrated, it is responsible for, among other things, the colours in betta fish. In addition, investigators showed that genetic variation in the BCO2gene influences the colours of feathers and skin in several bird species.

These genetic investigations have also been conducted in human eyes, where carotenoids accumulate in distinct patterns. BCO2 expressionis high in the peripheral retina,where there are no carotenoids, and low in the central fovea,where the carotenoids accumulate. This is reflected in single-cell sequencing.

The research into cone photoreceptors showed some surprising findings. The photoreceptors are rich in mitochondria, which is surprising because they rely on glycolysis rather than respiration mediated by these powerhouses of the cell. 

“Based on our research, we propose that photoreceptors are rich in mitochondria because, in the peripheral retina, BCO2-dependent carotenoid breakdown takes place in these organelles. In the central retina, the carotenoids are amassed in the mitochondria because of the absence of BCO2,” Dr von Lintig said.

However, an unresolved question is how the carotenoids enter the mitochondria. Dr von Lintig pointed out that carotenoids are lipids and cannot simply diffuse in an aqueous environment as sugars or amino acids do. They need specific transport proteins to move from one membrane to the other.

Aster-B

Dr von Lintig looked for candidate transport proteins that are expressed in the photoreceptors and found that the one with the highest expression in photoreceptors is Aster-B. “Aster-B was recently characterised in the context of cholesterol metabolism and transports cholesterol to mitochondria for steroid hormone production in the adrenal gland,” Dr von Lintig said. The next question addressed was whether Aster-B was really expressed in the retina.

To answer this question, the investigators analysed eye samples to determine whether Aster-B helped the carotenoids access the mitochondria. Interestingly, opposite expression patterns of BCO2 and Aster-B were found in the retina of human donor eyes. Immunohistochemistry showed Aster-B expression in the inner and outer segments of the photoreceptors.

Dr von Lintig also reported that Aster-B proteins, which constitute a novel family of lipid transfer proteins, have a domain-like structure. The proteins have a transmembrane domain in the endoplasmic reticulum, a lipid-binding domain and a phosphatidylserine-binding domain that connects them to another membrane. Aster-B proteins resemble bridges that catalyse the movement of lipids, such as cholesterol, from one membrane tothe other.

To determine whether the Aster proteins perform the same task with carotenoids, Dr von Lintig and colleagues invented a novel test system for which they bioengineered a new strain of bacteria that produced carotenoids. They expected that recombinant Aster proteins should extract the pigments from membranes when expressed in this bacterial strain. 

“That is exactly what they did,” he said. “Molecular modelling showed that the bulky carotenoid molecules fit perfectly into the binding cavity of Aster proteins.”

The investigators concluded that BCO2, which metabolises carotenoids and expresses in the peripheral retina, and Aster-B, which transports proteins, are important players in macula pigment metabolism and are responsible for the distinct pattern of accumulation of the pigments in the human retina.

Johannes von Lintig, PhD

E: Johannes.vonlintig@case.edu

This article is adapted from Dr von Lintig’s presentation at Cleveland Eye Bank Foundation’s 2022 Virtual Vision Research Symposium. He has no financial interest in this subject matter.