While the mammalian retina is well understood at the anatomical and physiological levels, little is known about the mechanisms that give rise to the retina's highly ordered pattern or its diverse neuronal cell types. Previous investigations have shown that gene disruption of the POU-IV class transcription factor Brn-3b (Brn-3.2) resulted in the loss of most retinal ganglion cells in retinas of postnatal mice. Here, we used lacZ and human placental alkaline phosphatase genes knocked into the brn-3b locus to follow the fate of brn-3b-mutant cells in the developing retina. We found that Brn-3b was not required for the initial commitment of retinal ganglion cell fate or for the migration of ganglion cells to the ganglion cell layer. However, Brn-3b was essential for the normal differentiation of retinal ganglion cells; without it, the cells underwent enhanced apoptosis. Retinal ganglion cells lacking brn-3b extended processes at the appropriate time in development, but these processes were disorganized, resulting in a thinner optic nerve. Explanted retinas from brn-3b-null embryos also extended processes when cultured in vitro, but the processes were shorter and less bundled than in wild-type retinas. Ultrastructural and marker analyses showed that the processes of mutant ganglion cells had dendritic rather than axonal features, suggesting that mutant cells formed dendrites in place of axons. These results suggest that Brn-3b regulates the activity of genes whose products play essential roles in the formation of retinal ganglion cell axons. (+info)
(2/78) Autoregulatory sequences are revealed by complex stability screening of the mouse brn-3.0 locus.
The POU-IV or Brn-3 class of transcription factors exhibit conserved structure, DNA-binding properties, and expression in specific subclasses of neurons across widely diverged species. In the mouse CNS, Brn-3.0 expression characterizes specific neurons from neurogenesis through the life of the cell. This irreversible activation of expression suggests positive autoregulation. To search for cis-acting elements that could mediate autoregulation we used a novel method, complex stability screening, which we applied to rapidly identify functional Brn-3.0 recognition sites within a large genomic region encompassing the mouse brn-3.0 locus. This method is based on the observation that the kinetic stability of Brn-3.0 complexes with specific DNA sequences, as measured by their dissociation half-lives, is highly correlated with the ability of those sequences to mediate transcriptional activation by Brn-3.0. The principal Brn-3.0 autoregulatory region lies approximately 5 kb upstream from the Brn-3.0 transcription start site and contains multiple Brn-3.0-binding sites that strongly resemble the optimal binding site for this protein class. This region also mediates transactivation by the closely related protein Brn-3.2, suggesting a regulatory cascade of POU proteins in specific neurons in which Brn-3.2 expression precedes Brn-3.0. (+info)
(3/78) The Brn-3b POU family transcription factor represses expression of the BRCA-1 anti-oncogene in breast cancer cells.
The BRCA-1 tumour supressor gene was identified on the basis of mutations which occur in familial breast cancer indicating that its inactivation can cause this disease. Although BRCA-1 does not appear to be mutated in sporadic breast cancer, its expression has been shown to be reduced in tumour material from such cases. We show here that mammary tumours which have reduced levels of BRCA-1 expression show enhanced expression of the Brn-3b POU family transcription factor at both the mRNA and protein levels. This elevated expression of Brn-3b is not found in normal mammary cells, benign tumours or in malignant tumour samples which do not exhibit reduced levels of BRCA-1. In contrast, no correlation was noted between BRCA-1 and expression of the related factor Brn-3a. Moreover, Brn-3b but not Brn-3a can strongly repress the BRCA-1 promoter approximately 20-fold in mammary tumour cells. To our knowledge, this is the first report of a transcription factor which regulates BRCA-1 expression. Thus, Brn-3b may play an important role in regulating expression of BRCA-1 in mammary tumours with enhanced expression of Brn-3b resulting in reduced BRCA-1 expression and thereby being potentially important in tumour development. (+info)
(4/78) All Brn3 genes can promote retinal ganglion cell differentiation in the chick.
Targeted gene disruption studies in the mouse have demonstrated crucial roles for the Brn3 POU domain transcription factor genes, Brn3a, Brn3b, Brn3c (now called Pou4f1, Pou4f2, Pou4f3, respectively) in sensorineural development and survival. During mouse retinogenesis, the Brn3b gene is expressed in a large set of postmitotic ganglion cell precursors and is required for their early and terminal differentiation. In contrast, the Brn3a and Brn3c genes, which are expressed later in ganglion cells, appear to be dispensable for ganglion cell development. To understand the mechanism that causes the functional differences of Brn3 genes in retinal development, we employed a gain-of-function approach in the chick embryo. We find that Brn3b(l) and Brn3b(s), the two isoforms encoded by the Brn3b gene, as well as Brn3a and Brn3c all have similar DNA-binding and transactivating activities. We further find that the POU domain is minimally required for these activities. Consequently, we show that all these Brn3 proteins have a similar ability to promote development of ganglion cells when ectopically expressed in retinal progenitors. During chick retinogenesis, cBrn3c instead of cBrn3b exhibits a spatial and temporal expression pattern characteristic of ganglion cell genesis and its misexpression can also increase ganglion cell production. Based on these data, we propose that all Brn3 factors are capable of promoting retinal ganglion cell development, and that this potential may be limited by the order of expression in vivo. (+info)
(5/78) The BRN-3A transcription factor protects sensory but not sympathetic neurons from programmed cell death/apoptosis.
Inactivation of the gene encoding the POU domain transcription factor BRN-3A results in the absence of specific neurons in knockout mice. Here we demonstrate for the first time a direct effect of BRN-3A on the survival of neuronal cells. Specifically, overexpression of BRN-3A in cultured trigeminal ganglion or dorsal root ganglion sensory neurons enhanced their survival following the withdrawal of nerve growth factor. Moreover, reduction of BRN-3A levels impaired the survival of these neurons. The survival of sympathetic neurons was not affected by either approach. Similarly, overexpression of BRN-3A activated the endogenous Bcl-2 gene in trigeminal neurons, but not in sympathetic neurons. The protective effect of BRN-3A on trigeminal neuron survival following nerve growth factor withdrawal significantly increased during embryonic development. In contrast, overexpression of the related factor BRN-3B enhanced survival of trigeminal neurons only at an early stage of embryonic development. Thus, BRN-3A (and in some circumstances, BRN-3B) can promote the survival of nerve growth factor-dependent sensory but not sympathetic neurons, allowing it to play a direct role in the survival of some (but not all) neuronal populations in the developing and adult nervous systems. (+info)
(6/78) Requirement for math5 in the development of retinal ganglion cells.
math5 is a murine orthologue of atonal, a bHLH proneural gene essential for the formation of photoreceptors and chordotonal organs in Drosophila. The expression of math5 coincides with the onset of retinal ganglion cell (RGC) differentiation. Targeted deletion of math5 blocks the initial differentiation of 80% of RGCs and results in an increase in differentiated amacrine cells. Furthermore, the absence of math5 abolishes the retinal expression of brn-3b and the formation of virtually all brn-3b-expressing RGCs. These results imply that math5 is a proneural gene essential for RGC differentiation and that math5 acts upstream to activate brn-3b-dependent differentiation processes in RGCs. (+info)
(7/78) A POU domain transcription factor-dependent program regulates axon pathfinding in the vertebrate visual system.
Axon pathfinding relies on the ability of the growth cone to detect and interpret guidance cues and to modulate cytoskeletal changes in response to these signals. We report that the murine POU domain transcription factor Brn-3.2 regulates pathfinding in retinal ganglion cell (RGC) axons at multiple points along their pathways and the establishment of topographic order in the superior colliculus. Using representational difference analysis, we identified Brn-3.2 gene targets likely to act on axon guidance at the levels of transcription, cell-cell interaction, and signal transduction, including the actin-binding LIM domain protein abLIM. We present evidence that abLIM plays a crucial role in RGC axon pathfinding, sharing functional similarity with its C. elegans homolog, UNC-115. Our findings provide insights into a Brn-3.2-directed hierarchical program linking signaling events to cytoskeletal changes required for axon pathfinding. (+info)
(8/78) The Ath5 proneural genes function upstream of Brn3 POU domain transcription factor genes to promote retinal ganglion cell development.
During retinogenesis, the Xenopus basic helix-loop-helix transcription factor Xath5 has been shown to promote a ganglion cell fate. In the developing mouse and chicken retinas, gene targeting and overexpression studies have demonstrated critical roles for the Brn3 POU domain transcription factor genes in the promotion of ganglion cell differentiation. However, the genetic relationship between Ath5 and Brn3 genes is unknown. To understand the genetic regulatory network(s) that controls retinal ganglion cell development, we analyzed the relationship between Ath5 and Brn3 genes by using a gain-of-function approach in the chicken embryo. We found that during retinogenesis, the chicken Ath5 gene (Cath5) is expressed in retinal progenitors and in differentiating ganglion cells but is absent in terminally differentiated ganglion cells. Forced expression of both Cath5 and the mouse Ath5 gene (Math5) in retinal progenitors activates the expression of cBrn3c following central-to-peripheral and temporal-to-nasal gradients. As a result, similar to the Xath5 protein, both Cath5 and Math5 proteins have the ability to promote the development of ganglion cells. Moreover, we found that forced expression of all three Brn3 genes also can stimulate the expression of cBrn3c. We further found that Ath5 and Brn3 proteins are capable of transactivating a Brn3b promoter. Thus, these data suggest that the expression of cBrn3c in the chicken and Brn3b in the mouse is initially activated by Ath5 factors in newly generated ganglion cells and later maintained by a feedback loop of Brn3 factors in the differentiated ganglion cells. (+info)