Role of the transketolase-like-1 gene in the development of the modern human brain

Role of the transketolase-like-1 gene in the development of the modern human brain

In a recently published study in Science Researchers showed how transketolase-like 1 (TKTL1) gene expression influences neocortical neurogenesis in modern humans.

Lernen: Menschliches TKTL1 impliziert eine größere Neurogenese im frontalen Neokortex moderner Menschen als bei Neandertalern. Bildnachweis: iurii/Shutterstock

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Interestingly, a single amino acid substitution in TKTL1, from lysine in monkeys and archaic humans to arginine in modern humans, altered the function and organization of brain layers such as the neocortex, particularly in the frontal lobe. The evolutionary changes in the brain's neocortex and the concomitant increase in neuron production improved the cognitive abilities of modern humans. Endocast analyzes have shown that the endocranial volume of modern humans and Neanderthals was similar, suggesting similar size of the neocortex. However, fossils could not help assess whether similar neocortex size implied similar neocortical neuron production.

A previous study by Pinson et al. analyzed the effect of a single amino acid change in the TKTL1 protein on the production of basal radial glia (bRG), the cells that form most of the neocortex. They observed that in organoids, the human variant of TKTL1 (hTKTL1) generated more neuroprogenitors than the archaic variant (aTKTL1) found in Neanderthals, Denisovans and other primates. Maybe that's why modern humans have more neocortex in their brains than Neanderthals.

About studying

In the present study, researchers examined the role of TKTL1 in the development of the neocortex, which in turn influences the number of neuroprogenitors. In addition, they determined whether aTKTL1 and hTKTL1 have similar effects on neuroprogenitors during neocortex development.

The study used gene overexpression in developing mouse and ferret neocortex. Likewise, they used knockout technology in fetal human neocortical tissue and genome editing technology to study cerebral organoids.

Using clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR-associated protein 9 (Cas9)-mediated hTKTL1 knockout technology, they showed that hTKTL1 knockout in fetal human neocortical tissue significantly reduced brG numbers. Next, the team used cerebral organoids to validate their results. These mini-brain structures made from human embryonic stem cells had aTKTL1 instead of hTKTL1.

Specifically, the team obtained total ribonucleic acid (RNA) from mock and gene-edited H9-derived human organoids. They used quantitative real-time PCR (qPCR) to confirm that these organoids expressed TKTL1 messenger RNA (mRNA), followed by Sanger sequencing of the PCR products. They mapped mock and gene-edited organoids to the human TKTL1 gene to confirm the expression of hTKTL1 mRNA in the mock-edited human organoids and archaeal TKTL1 mRNA in the gene-edited human organoids.

Before immunofluorescence, they cut coronal cryosections with a thickness of 20–50 μm using a cryostat and stored these samples at −20 °C. First, the team subjected a single cell suspension from the embryonic mouse neocortex to cell surface staining. Next, they microdissected it under an epifluorescence stereomicroscope. In this way, they obtained the electroporated region of the neocortex. Finally, the team used mass spectrometry (MS) to determine the concentrations of acetyl-coenzyme A (acetyl-CoA) in the bRG isolated by fluorescence-activated cell sorting (FACS).

Study results

Of course, the mouse embryo neocortex lacks TKTL1 expression. Introduction of the hTKTL1 gene into the neocortex of mouse embryos increased the frequency of bRG without affecting the basal intermediate progenitors (bIPs) and apical progenitor population, which in turn increased cortical neuron production over time, particularly of late developing upper layer neurons. Conversely, aTKTL1, which differed by only one amino acid, did not increase bRG abundance.

Furthermore, the authors found that hTKTL1 expression increased bRG abundance and the proportion of multiprocess bRG, a hallmark of bRG that can be self-reinforcing, in developing ferret neocortex. Consequently, the ferret's gyrus size increased.

In addition, the researchers observed that the aTKTL1-expressing cell organoids contained fewer bRG and neuroprogenitor cells. This finding confirmed that the lysine-to-arginine substitution in hTKTL1 was essential for the maintenance of the bRG population in this human brain model. Likewise, hTKTL1 expression increased in neuroprogenitors during neurogenesis in the human fetal neocortex. Interestingly, this expression was higher in the developing frontal lobe than in the developing occipital lobe.

Another interesting observation was that hTKTL1 enhanced the bRG population through two sequential metabolic pathways, the pentose phosphate pathway (PPP) and fatty acid synthesis. Therefore, when the researchers suppressed the PPP or fatty acid synthesis pathways with a series of specific inhibitors, the hTKTL1-induced increase in the bRG population in the embryonic mouse neocortex ceased. Similarly, bRG numbers were reduced in human fetal neocortical tissue.

Furthermore, the authors found that hTKTL1-stimulated bRGs, but not aTKTL1-induced bRGs, had a higher concentration of acetyl-CoA, a metabolite crucial for fatty acid synthesis. Thus, only hTKTL1 promoted the synthesis of membrane lipids containing a type of fatty acid required for the outgrowth of bRG processes, increasing their population.

Conclusions

The current study showed what differentiates neocortical neurogenesis in modern humans from Neanderthals. Although these archaic humans had similar brains (in size) to modern humans, a single lysine-to-arginine substitution in the hTKTL1 gene resulted in bRG abundance but not that of bIPs, which in turn generated more neocortical neurons in modern humans. Furthermore, the researchers showed that the hTKTL1 effect required PPP and fatty acid synthesis, and inhibition of these pathways reduced bRG abundance in fetal human neocortical tissue.

Reference:

• Anneline Pinson, Lei Xing, Takashi Namba, Nereo Kalebic, Jula Peters, Christina Eugster Oegema, Sofia Traikov, Katrin Reppe, Stephan Riesenberg, Tomislav Maricic, Razvan Derihaci, Pauline Wimberger, Svante Pääbo, Wieland B. Huttner. (2022). Menschliches TKTL1 impliziert eine größere Neurogenese im frontalen Neokortex moderner Menschen als bei Neandertalern. Wissenschaft. doi: 10.1126/science.abl6422 https://www.science.org/doi/10.1126/science.abl6422

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