Journal Club: Reproductive Biology, Immunology, Neuroscience, and Genome Engineering.

Graduate Division

REPRODUCTIVE BIOLOGY: Mammalian Fused is essential for sperm head shaping and periaxonemal structure formation during spermatogenesis. Nozawa, I., et al. (Chuang). Dev Biol. 2014 Feb. 10. Epub ahead of print.

As depicted in many middle school sexual education videos, sperm rely on their vigorously beating tail to propel themselves on their long journey to the egg. The development of this flagellum is part of the final phase of spermatogenesis.

Although the sequence of events occurring during this final metamorphosis is well described, the precise mechanism remains poorly understood.

In this paper, Nozawa and colleagues have demonstrated an essential role for the protein Fused in this process. They find that germ cell-specific deletion of Fused rendered male mice infertile. They further showed that Fused localizes to microtubule-organizing structures. In the absence of Fused, the axoneme, the core flagellar structure, is present but problems in the bordering periaxonemal structure lead to poor motility.  

IMMUNOLOGY: PKCδ promotes transitional B cell negative selection and limits proximal BCR signaling to enforce tolerance. Limnander, A., et al. (Roose). Mol Cell Biol. 2014 Feb. 10. Epub ahead of print.

As B cells develop, genetic recombination produces highly variable B cell receptors. This diversity results in being ready to produce antibodies against a nearly limitless range of potential targets, but also invariably includes receptors that recognize components of the human body.

Several mechanisms normally act to eliminate or control these self-reactive B cells. Disruptions in these checks can result in severe autoimmunity, as is seen in rare patients with mutations in the signaling cascade kinase PKCδ.

In this article, the authors investigated how loss of PKCδ leads to autoimmunity. They showed that this protein is necessary for activation of a signaling cascade that results in the death of self-reactive B cells. They also found that it normally dampens signaling from the B cell receptor, so loss of its function makes the B cells aberrantly easy to activate.

NEUROSCIENCE: Olig1 function is required to repress dlx1/2 and interneuron production in mammalian brain. Silbereis, J.C., et al. (Rowitch). Neuron. 2014. 81(3):574-87.

The healthy cerebral cortex requires a balance between excitatory and stimulatory signals—too much of either has been associated with neural disorders such as epilepsy.

The appropriate relative numbers of excitatory pyramidal neurons and inhibitory interneurons are established during embryonic development in a carefully regulated process. Previous research has shown that normal development of inhibitory interneurons requires the action of the transcription factors Dlx1/2, but the detailed mechanism has remained unclear.

The authors here addressed this question by examining the role of the transcription factor Olig1. They found that deletion of Olig1 in mice results in a 30 percent increase in cortical interneurons because Olig1 usually represses the transcription factor Dlx1/2. The researchers suggested that Olig1 merits investigation as a potential therapeutic target. 

GENOME ENGINEERING: Isolation of single-base genome-edited human iPS cells without antibiotic selection. Miyaoka, Y., et al. (Conklin). Nat Methods. 2014 Feb. 9. Epub ahead of print.

Many human genetic disorders are due to point mutations, in which only a single amino acid is altered. When introducing these mutations into cells for study, scientists usually use antibiotics to select for successfully altered cells, which requires genomic changes in addition to the point mutation. 

In this paper, the authors described a method for efficiently identifying rare mutations, making antibiotic selection unnecessary, and demonstrated it by introducing specific point mutations into five different genes.

The method relies on highly sensitive detection of mutants and progressive enrichments. Cells are plated into multiple wells, some of which will by chance have a higher proportion of cells with the desired mutation. The most enriched well can be plated again into several wells, the most enriched of these selected, and so forth. After several rounds, the researchers isolated clones initially present at a frequency of only one in two thousand.