a-fast-quantitative-method-for-post-translational-modification

A Fast and Quantitative Method for Post-translational Modification and Variant Enabled Mapping of Peptides to Genomes

Cross-talk between genes, transcripts, and proteins is the key to cellular responses; hence, analysis of molecular levels as distinct entities is slowly ...

Boston Children's Hospital, Harvard Medical School

Neurons differ in their responses to injury, but the underlying mechanisms remain poorly understood. Using quantitative proteomics, we characterized the injury-triggered response from purified intact and axotomized retinal ganglion cells (RGCs). Subsequent informatics analyses revealed a network of injury-response signaling hubs. In addition to confirming known players, such as mTOR, this also identified new candidates, such as c-myc, NFκB, and Huntingtin. Similar to mTOR, c-myc has been implicated as a key regulator of anabolic metabolism and is downregulated by axotomy. Forced expression of c-myc in RGCs, either before or after injury, promotes dramatic RGC survival and axon regeneration after optic nerve injury. Finally, in contrast to RGCs, neither c-myc nor mTOR was downregulated in injured peripheral sensory neurons. Our studies suggest that c-myc and other injury-responsive pathways are critical to the intrinsic regenerative mechanisms and might represent a novel target for developing neural repair strategies in adults.

Injury-induced decline of intrinsic regenerative ability revealed by quantitative proteomics.

After axotomy, neuronal survival and growth cone re-formation are required for axon regeneration. We discovered that doublecortin-like kinases (DCLKs), members of the doublecortin (DCX) family expressed in adult retinal ganglion cells (RGCs), play critical roles in both processes, through distinct mechanisms. Overexpression of DCLK2 accelerated growth cone re-formation in vitro and enhanced the initiation and elongation of axon re-growth after optic nerve injury. These effects depended on both the microtubule (MT)-binding domain and the serine-proline-rich (S/P-rich) region of DCXs in-cis in the same molecules. While the MT-binding domain is known to stabilize MT structures, we show that the S/P-rich region prevents F-actin destabilization in injured axon stumps. Additionally, while DCXs synergize with mTOR to stimulate axon regeneration, alone they can promote neuronal survival possibly by regulating the retrograde propagation of injury signals. Multifunctional DCXs thus represent potential targets for promoting both survival and regeneration of injured neurons.

Doublecortin-Like Kinases Promote Neuronal Survival and Induce Growth Cone Reformation via Distinct Mechanisms.

The ability to integrate 'omics' (i.e. transcriptomics and proteomics) is becoming increasingly important to the understanding of regulatory mechanisms. There are currently no tools available to identify differentially expressed genes (DEGs) across different 'omics' data types or multi-dimensional data including time courses. We present fCI (f-divergence Cut-out Index), a model capable of simultaneously identifying DEGs from continuous and discrete transcriptomic, proteomic and integrated proteogenomic data. We show that fCI can be used across multiple diverse sets of data and can unambiguously find genes that show functional modulation, developmental changes or misregulation. Applying fCI to several proteogenomics datasets, we identified a number of important genes that showed distinctive regulation patterns. The package fCI is available at R Bioconductor and http://software.steenlab.org/fCI/.

Judith A.J. Steen

Department of Neurobiology,
F. M. Kirby Neurobiology Center,
Department of Neurobiology, F. M. Kirby Neurobiology Center

Boston Children's Hospital, Harvard Medical School

A Fast and Quantitative Method for Post-translational Modification and Variant Enabled Mapping of Peptides to Genomes

Judith A.J. Steen

Department of Neurobiology,F. M. Kirby Neurobiology Center,Department of Neurobiology, F. M. Kirby Neurobiology Center

Boston Children's Hospital, Harvard Medical School

A Fast and Quantitative Method for Post-translational Modification and Variant Enabled Mapping of Peptides to Genomes

Department of Neurobiology,
F. M. Kirby Neurobiology Center,
Department of Neurobiology, F. M. Kirby Neurobiology Center