Post by Admin on Aug 3, 2022 18:56:36 GMT
Longer metaphase and fewer chromosome segregation errors in modern human than Neanderthal brain development
Abstract
Since the ancestors of modern humans separated from those of Neanderthals, around 100 amino acid substitutions spread to essentially all modern humans. The biological significance of these changes is largely unknown. Here, we examine all six such amino acid substitutions in three proteins known to have key roles in kinetochore function and chromosome segregation and to be highly expressed in the stem cells of the developing neocortex. When we introduce these modern human-specific substitutions in mice, three substitutions in two of these proteins, KIF18a and KNL1, cause metaphase prolongation and fewer chromosome segregation errors in apical progenitors of the developing neocortex. Conversely, the ancestral substitutions cause shorter metaphase length and more chromosome segregation errors in human brain organoids, similar to what we find in chimpanzee organoids. These results imply that the fidelity of chromosome segregation during neocortex development improved in modern humans after their divergence from Neanderthals.
INTRODUCTION
The neocortex is unique to mammals and the seat of sensory and motor activities (1). During the evolution of humans, the neocortex increased drastically in size. This is widely considered to be associated with the development of human cognitive abilities (2–8). Quantitative changes that lead to an increase in neocortex size include, for example, increases in the proliferative capacity and numbers of neocortical stem and progenitor cells, and consequently in the numbers of neurons and macroglial cells generated by them (3–8). Comparatively less is known about qualitative changes in neocortex development during hominin evolution that may have occurred concomitant with the increase in neocortex size. However, substitutions and duplications affecting the genes FOXP2 (9, 10) and SRGAP2C (11, 12) have been shown to affect synapse formation and connectivity, resulting in improved learning in mouse models (13, 14).
Brain organoids are useful tissue models for neural progenitors, especially for those in the ventricular zone (15–17). We previously compared the mitotic behavior of neocortical stem and progenitor cells in humans, chimpanzees, and orangutans, using induced pluripotent stem cell (iPSC)–derived cerebral organoids (18). We found that human proliferating apical progenitors (APs), the cells that line the ventricles and from which all other neural cells in the developing neocortex originate, spend around 50% more time in mitotic metaphase than the APs of chimpanzees and orangutans. Metaphase is the step in mitosis where the cell finalizes the preparations to start the segregation and equal distribution of the chromosomes to the two daughter cells (19). Hence, these differences in metaphase length raise the possibility that the fidelity of chromosome segregation during AP mitosis might differ between humans and apes, with potential consequences for neocortex development and function.
We focus on the roles of three proteins KIF18a [also known as (a.k.a.) Kinesin 8], KNL1 (a.k.a. CASC5), and SPAG5 (a.k.a. astrin), which are highly expressed in the germinal zones of the developing neocortex and are associated with mitotic spindle, kinetochore, and chromosome segregation functions. The kinetochore is a complex, three-dimensional (3D), multiprotein structure mediating the attachment of chromosome centromeres with the ends of kinetochore microtubules (20–22). An important role of kinetochores is to facilitate spindle assembly checkpoint (SAC) function, which regulates the onset of chromosome segregation when chromosomes are correctly aligned at the metaphase plate (23–26). The three proteins stand out because they carry amino acid substitutions found in all present-day humans but are essentially absent in apes and in Neanderthals or Denisovans, i.e., so-called archaic humans, which separated from the evolutionary lineage leading to modern humans about half a million years ago (27). Any functional consequences of these substitutions would thus be unique to modern humans (28–30).
KIF18a, which carries one modern human-specific amino acid substitution, is a motor protein of the kinesin family that is involved in regulating correct chromosome positioning and attachment to kinetochore microtubules, and their bi-orientation within the mitotic spindle (31–33). KNL1, which carries two modern human-specific amino acid substitutions, is part of the outer kinetochore, which is required for attachment of the kinetochores to the microtubules. It is also a main docking site for key proteins of the SAC, such as BubR1 and Mad1, and therefore important for chromosome alignment and segregation (34–37). SPAG5, which carries three modern human-specific amino acid substitutions, is a microtubule-associated protein recruited to kinetochores and is important for the stability of attachment of kinetochores to microtubules (38–41).
We show here that APs in the embryonic neocortex of mice where the modern human substitutions in KIF18a and KNL1 have been introduced by genome editing exhibit longer metaphases, more SAC-positive kinetochores, and fewer mis-segregating chromosomes. In converse experiments, where human embryonic stem cells (ESCs) that carry the ancestral variants of KIF18a and KNL1 are used to generate cerebral organoids, shorter metaphases, less SAC-positive kinetochores, and more chromosome segregation defects are observed in APs. Together, our data suggest that the three amino acid substitutions in KIF18a and KNL1 cause fewer chromosome inheritance errors to occur in APs of modern humans than in archaic humans and apes.
Abstract
Since the ancestors of modern humans separated from those of Neanderthals, around 100 amino acid substitutions spread to essentially all modern humans. The biological significance of these changes is largely unknown. Here, we examine all six such amino acid substitutions in three proteins known to have key roles in kinetochore function and chromosome segregation and to be highly expressed in the stem cells of the developing neocortex. When we introduce these modern human-specific substitutions in mice, three substitutions in two of these proteins, KIF18a and KNL1, cause metaphase prolongation and fewer chromosome segregation errors in apical progenitors of the developing neocortex. Conversely, the ancestral substitutions cause shorter metaphase length and more chromosome segregation errors in human brain organoids, similar to what we find in chimpanzee organoids. These results imply that the fidelity of chromosome segregation during neocortex development improved in modern humans after their divergence from Neanderthals.
INTRODUCTION
The neocortex is unique to mammals and the seat of sensory and motor activities (1). During the evolution of humans, the neocortex increased drastically in size. This is widely considered to be associated with the development of human cognitive abilities (2–8). Quantitative changes that lead to an increase in neocortex size include, for example, increases in the proliferative capacity and numbers of neocortical stem and progenitor cells, and consequently in the numbers of neurons and macroglial cells generated by them (3–8). Comparatively less is known about qualitative changes in neocortex development during hominin evolution that may have occurred concomitant with the increase in neocortex size. However, substitutions and duplications affecting the genes FOXP2 (9, 10) and SRGAP2C (11, 12) have been shown to affect synapse formation and connectivity, resulting in improved learning in mouse models (13, 14).
Brain organoids are useful tissue models for neural progenitors, especially for those in the ventricular zone (15–17). We previously compared the mitotic behavior of neocortical stem and progenitor cells in humans, chimpanzees, and orangutans, using induced pluripotent stem cell (iPSC)–derived cerebral organoids (18). We found that human proliferating apical progenitors (APs), the cells that line the ventricles and from which all other neural cells in the developing neocortex originate, spend around 50% more time in mitotic metaphase than the APs of chimpanzees and orangutans. Metaphase is the step in mitosis where the cell finalizes the preparations to start the segregation and equal distribution of the chromosomes to the two daughter cells (19). Hence, these differences in metaphase length raise the possibility that the fidelity of chromosome segregation during AP mitosis might differ between humans and apes, with potential consequences for neocortex development and function.
We focus on the roles of three proteins KIF18a [also known as (a.k.a.) Kinesin 8], KNL1 (a.k.a. CASC5), and SPAG5 (a.k.a. astrin), which are highly expressed in the germinal zones of the developing neocortex and are associated with mitotic spindle, kinetochore, and chromosome segregation functions. The kinetochore is a complex, three-dimensional (3D), multiprotein structure mediating the attachment of chromosome centromeres with the ends of kinetochore microtubules (20–22). An important role of kinetochores is to facilitate spindle assembly checkpoint (SAC) function, which regulates the onset of chromosome segregation when chromosomes are correctly aligned at the metaphase plate (23–26). The three proteins stand out because they carry amino acid substitutions found in all present-day humans but are essentially absent in apes and in Neanderthals or Denisovans, i.e., so-called archaic humans, which separated from the evolutionary lineage leading to modern humans about half a million years ago (27). Any functional consequences of these substitutions would thus be unique to modern humans (28–30).
KIF18a, which carries one modern human-specific amino acid substitution, is a motor protein of the kinesin family that is involved in regulating correct chromosome positioning and attachment to kinetochore microtubules, and their bi-orientation within the mitotic spindle (31–33). KNL1, which carries two modern human-specific amino acid substitutions, is part of the outer kinetochore, which is required for attachment of the kinetochores to the microtubules. It is also a main docking site for key proteins of the SAC, such as BubR1 and Mad1, and therefore important for chromosome alignment and segregation (34–37). SPAG5, which carries three modern human-specific amino acid substitutions, is a microtubule-associated protein recruited to kinetochores and is important for the stability of attachment of kinetochores to microtubules (38–41).
We show here that APs in the embryonic neocortex of mice where the modern human substitutions in KIF18a and KNL1 have been introduced by genome editing exhibit longer metaphases, more SAC-positive kinetochores, and fewer mis-segregating chromosomes. In converse experiments, where human embryonic stem cells (ESCs) that carry the ancestral variants of KIF18a and KNL1 are used to generate cerebral organoids, shorter metaphases, less SAC-positive kinetochores, and more chromosome segregation defects are observed in APs. Together, our data suggest that the three amino acid substitutions in KIF18a and KNL1 cause fewer chromosome inheritance errors to occur in APs of modern humans than in archaic humans and apes.