Felix Karim

KSR, a novel protein kinase required for RAS signal transduction

We have identified and characterized two genes in Drosophila whose products are required for activated RAS to signal with normal efficiency, but do not appear to effect signaling by activated RAF. One encodes the beta subunit of type I geranylgeranyl transferase, a prenylation enzyme essential for targeting RAS to the plasma membrane. The other encodes a protein kinase that we have named kinase suppressor of ras (ksr). By genetic criteria, we show that KSR functions in multiple receptor tyrosine kinase pathways. We have isolated mammalian homologs of KSR that, together with the Drosophila gene, define a novel class of kinases. Our results suggest that KSR is a general and evolutionarily conserved component of the RAS signaling pathway that acts between RAS and RAF.

Temporal coordination of regulatory gene expression by the steroid hormone ecdysone.

In Drosophila, pulses of the steroid hormone ecdysone function as temporal signals that trigger the major postembryonic developmental transitions. The best characterized of these pulses activates a series of puffs in the polytene chromosomes as it triggers metamorphosis. A small set of early puffs is induced as a primary response to the hormone. These puffs encode regulatory proteins that both repress their own expression and activate a large set of late secondary response genes. We have used Northern blot analysis of RNA isolated from staged animals and cultured organs to study the transcription of three primary response regulatory genes, E75, BR‐C and EcR. Remarkably, their patterns of transcription in late larvae can be defined in terms of two responses to different ecdysone concentrations. The class I transcripts (E74B and EcR) are induced in mid‐third instar larvae in response to the low, but increasing, titer of ecdysone. As the hormone concentration peaks in late third instar larvae, these transcripts are repressed and the class II RNAs (E74A, E75A and E75B) are induced. The BR‐C RNAs appear to have both class I and class II characteristics. These data demonstrate that the relatively simple profile of a hormone pulse contains critical temporal information that is transduced into waves of primary response regulatory gene activity.

phyllopod functions in the fate determination of a subset of photoreceptors in drosophila

phyllopod (phyl) encodes a novel protein required for fate determination of photoreceptors R1, R6, and R7, the last three photoreceptors to be recruited into the ommatidia of the developing Drosophila eye. Genetic data suggests that phyl acts downstream of Ras1, raf, and yan to promote neuronal differentiation in this subset of photoreceptors. Ectopic expression of phyl in the cone cell precursors mimics the effect of ectopic activation of Ras1, suggesting that phyl expression is regulated by Ras1. phyl is also required for embryonic nervous system and sensory bristle development.

PTP-ER, a Novel Tyrosine Phosphatase, Functions Downstream of Ras1 to Downregulate MAP Kinase during Drosophila Eye Development

Activation of ERK/MAPK is a key event downstream of RAS. The duration, extent, and timing of MAPK activity is integral to signal specificity. Consequently, inactivation of MAPK by phosphatases has emerged as a critical element in the precise control of signal output. We have cloned and characterized a novel cytoplasmic protein tyrosine phosphatase, PTP-ER, which is related to mammalian PCPTP1, LC-PTP/HePTP, and STEP tyrosine phosphatases. PTP-ER mutants produce extra R7 cells and enhance activated Ras1 signaling. Ectopic expression of PTP-ER dramatically inhibits RAS1/MAPK signaling. PTP-ER binds to and inactivates Drosophila ERK/MAPK; however, it is unable to dephosphorylate and downregulate Drosophila MAPKSevenmaker. Resistance to PTP-ER activity partially accounts for the Sevenmaker mutant phenotype.

Non-ATP-competitive kinase inhibitors - enhancing selectivity through new inhibition strategies.

Background: ATP-competitive inhibitors of protein kinases have been successfully developed for life-threatening indications such as cancer. However, owing to the similarity of the ATP binding sites between kinases, it has been challenging to identify specific inhibitors. The progress towards the generation of kinase inhibiting drugs for more chronic indications has been slowed by the concern that low specificity kinase inhibitors will have undesired toxicities. Objective and methods: We have reviewed the scientific and patent literature to summarize alternative strategies that are being used to develop non-ATP-competitive kinase inhibitors with greater selectivity. Results/conclusion: Several new approaches are being taken to achieve selectivity. Among these, the use of small peptide therapeutics is particularly promising and is already yielding drugs that are demonstrating promise in human clinical trials.