Scientific Understanding of Consciousness
Consciousness as an Emergent Property of Thalamocortical Activity

Corticospinal Development and Evolution



Nature 486, 74–79  (07 June 2012)

Cis-regulatory control of corticospinal system development and evolution

Department of Neurobiology and Kavli Institute for Neuroscience, Yale University School of Medicine, New Haven, Connecticut 06510, USA

Sungbo Shim, Kenneth Y. Kwan, Mingfeng Li & Nenad Šestan

Department of Cell Biology and Orthopaedic and Rheumatologic Research Center, Cleveland Clinic Lerner Research Institute, Cleveland, Ohio 44195, USA

Veronique Lefebvre


The co-emergence of a six-layered cerebral neocortex and its corticospinal output system is one of the evolutionary hallmarks of mammals. However, the genetic programs that underlie their development and evolution remain poorly understood. Here we identify a conserved non-exonic element (E4) that acts as a cortex-specific enhancer for the nearby gene Fezf2 (also known as Fezl and Zfp312), which is required for the specification of corticospinal neuron identity and connectivity. We find that SOX4 and SOX11 functionally compete with the repressor SOX5 in the transactivation of E4. We show evidence supporting the emergence of functional SOX-binding sites in E4 during tetrapod evolution, and their subsequent stabilization in mammals and possibly amniotes. These findings reveal that SOX transcription factors converge onto a cis-acting element of Fezf2 and form critical components of a regulatory network controlling the identity and connectivity of corticospinal neurons.

The emergence and expansion of the neocortex in mammals has been crucial to the evolution of complex perceptual, cognitive, emotional and motor abilities. The neocortex is organized into six layers based largely on the distinct subtypes of excitatory projection (or pyramidal) neurons and their patterns of connectivity. Upper-layer (L2, L3 and L4) projection neurons form synaptic connections solely with other cortical neurons. By contrast, the majority of neurons in the deeper layers (L5 and L6) project to subcortical regions. Studies of laminar inversion in reeler mice lacking the reelin (RELN) protein have shown that neuron identity and connectivity are determined by birth order rather than by laminar position, suggesting that neuronal specification and positioning are largely separately encoded.

The layer-specific pattern of connectivity is dependent on cortical areas. The long-range projections of L5 neurons in somatosensory-motor areas form the corticospinal (CS) system that directly connects the neocortex with various subcortical regions. A major component of the system, the CS (or pyramidal) tract, descends through the brainstem and into the spinal cord to provide a high degree of direct control over the precise motor functions affected in many clinical conditions. Despite these important functional implications, the genetic programs controlling CS system development and evolution remain unclear.

Phenotypic specification and evolution of neural circuits depend on precise regulation of the timing, location and level of gene expression. Transcriptional control via cis-regulatory elements has emerged as a crucial mechanism. The cis-regulatory mechanisms underlying the specification of distinct neuronal cell types and circuits, however, remain poorly understood. Specification of CS neurons and the formation of the CS tract critically depend on Fezf2, which encodes a zinc-finger transcription factor  highly enriched in early cortical progenitor cells and their deep-layer neuron progenies. Research findings indicate that the precisely regulated transcription of Fezf2 is probably critical to the proper specification of distinct types of cortical projection neurons.

In this study, we used bacterial artificial chromosome (BAC) engineering and genetic inactivation in mice to identify and characterize a cortex-specific Fezf2 enhancer and its trans-regulators. We show that three SOX transcription factors converge onto the Fezf2 enhancer to control CS system development via functional binding sites that emerged in tetrapods. We also found that Sox4 and Sox11 are required for Fezf2-independent regulation of cortical RELN expression and laminar organization. Thus, these findings reveal novel developmental genetic programs that control layer formation and CS neuron identity, and the regulatory mechanisms by which they may have evolved.

On the basis of the remarkable similarity in cortical expression pattern of Fezf2 between mouse and human, we proposed that the regulatory elements are also highly evolutionarily conserved.

These results demonstrate that E4 is a cis-regulatory module acting as a cortex-specific enhancer of Fezf2.

Our data provide critical insight into the genes and regulatory components controlling CS system development, centring on the cis-regulation of Fezf2 by SOX4 and SOX11.

Our results indicate that, following their emergence in tetrapods, functional SOX-binding sites have retained high conservation through purifying selection in mammals and some amniotes, thus directly linking species variations in regulatory sequences to functional outcomes. E4 sequence substitutions in SB2 may constitute an evolutionary turning point for Fezf2 function during forebrain development, possibly facilitating the formation of descending telencephalic pathways including the CS system. Whereas minor projections from the ventral (subpallial) telencephalon to the spinal cord are present in amphibians, direct dorsal telencephalo-spinal projections resembling the CS tract have been reported only in mammals and some birds. We propose that the concurrent emergence of the described regulatory mechanisms and direct telencephalo-spinal projections in early amniotes, together with subsequent changes in genetic programs driving the patterning and expansion of a six-layered dorsal pallium, made possible the evolution of the CS system in mammals.

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