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Page 221 Mejia et al. J Transl Genet Genom 2024;8:216-24 https://dx.doi.org/10.20517/jtgg.2024.11

[15]
alteration in the phosphorylation of PKCδ in BTHS lymphoblasts . Hence, altered phosphorylation of
PKCδ might contribute to an attenuated mitochondrial PKCδ signaling in these cells.

The molecular components that mediate PKCδ signaling in mitochondria are beginning to emerge.
Mitochondria contain a high molecular weight functional complex, which includes cytochrome c as the
upstream driver of PKCδ, and it uses the adapter protein p66Shc as the assembly platform with vitamin A
(retinol) [12,14,22] . All four partners are required for functional PKCδ signaling. BN-PAGE immunoblot analysis
of mitochondrial proteins not only has the advantage of probing for expression of individual proteins but
may additionally be used to detect if these proteins are associated with higher molecular weight complexes.
Using this approach, we observed a reduction in PKCδ associated with a higher molecular weight complex
in BTHS B lymphoblasts mitochondria. It is possible that the decreased association of PKCδ with the high
molecular weight complex was associated with accumulation of MLCL in our BTHS lymphoblasts and that
the increased ratio of MLCL to CL may affect inner membrane structural integrity such that the high
molecular weight complex dissociates. Previous studies have demonstrated that MLCL-protein interactions
compromise the stability of the protein-dense mitochondrial inner membrane .
[23]
The PKCδ/retinol complex signals the pyruvate dehydrogenase complex for enhanced flux of pyruvate into
the Krebs cycle [12,14] . Interestingly, in the UK BTHS NHS clinic, almost half of the BTHS boys examined
showed signs of Vitamin A deficiency (Nicol Clayton: https://www.youtube.com/watch?v=wNDr_oCTJ7A).
However, supplementation with Vitamin A did not increase plasma levels. This was not because tissue levels
were low but possibly due to increased levels of Vitamin A (retinyl esters) in chylomicrons. It is possible
that this unique observation is coupled to defective mitochondrial PKCδ signaling, which might contribute
to reduced ATP production in the Krebs cycle through alteration in the mitochondrial PKCδ/retinol
signaling complex and contribute to the multitude of bioenergetic defects observed in BTHS. However, it is
unknown whether the decreased association of PKCδ within the high molecular weight complex in Barth
Syndrome is a cause of mitochondrial dysfunction or an effect of mitochondrial dysfunction.

Creatine is an important energy metabolite that is used as an energy buffer. Impaired mitochondrial
oxidative phosphorylation, as seen in BTHS, may require increased energy demand from alternative sources
such as ATP synthesis from enhanced glucose uptake and oxidation or ATP resynthesis from ADP and
phosphocreatine. Interestingly, creatine supplementation has been shown to increase glucose uptake and
[24]
oxidation and adenosine monophosphate kinase (AMPK) phosphorylation in skeletal muscle cells . We
previously reported that increased AMPK phosphorylation and its activation accompanied elevated glucose
uptake in BTHS B lymphoblasts . Enhanced glucose uptake in TAFZZIN-deficient cells may additionally
[17]
be linked to the upregulation of pyruvate dehydrogenase 4 mediated through AMPK activation and
[25]
transcriptional upregulation by forkhead box protein O1 . Moreover, creatine kinase has been shown to be
mildly elevated in the plasma of some BTHS patients . Thus, enhanced creatine uptake might be required
[26]
to support phosphocreatine generation if creatine kinase was depleted in cells of these patients. In the
current study, creatine uptake was significantly enhanced in BTHS B lymphoblasts compared to controls.
Interestingly, the human creatine transporter (CRTR) gene was shown to be localized on Xq28 and, at one
[27]
time, was hypothesized to be a candidate gene for BTHS and infantile cardiomyopathy . However, a
subsequent study by Sylvia Bione identified the actual locus of the human TAFAZZIN gene to be
Xq28.12 . Whether creatine supplementation improves the health of BTHS patients is unknown. A
[28]
previous study indicated that creatine supplementation in humans improved performance during exercise
[29]
of high to maximal intensity .