Germline deletion reveals a non-essential role of the atypical MAPK6/ERK3

Abstract

words) 24MAPK6/ERK3 is an atypical member of the MAPKs. An essential role has been suggested by the 25 perinatal lethal phenotype of ERK3 knockout mice carrying a lacZ insertion in exon 2 due to 26 pulmonary disfunction and by defects in function, activation and positive selection of T cells. To 27 study the role of ERK3 in vivo, we generated mice carrying a conditional Erk3 allele with exon3 28 flanked by LoxP sites. Loss of ERK3 protein was validated after deletion of Erk3 in the female 29 germ line using zona pellucida 3 (Zp3)-cre and a clear reduction of the protein kinase MK5 is 30 detected, providing first evidence for the existence of the ERK3/MK5 signaling complex in vivo. 31In contrast to the previously reported Erk3 knockout phenotype, these mice are viable and 32 fertile, do not display pulmonary hypoplasia, acute respiratory failure, abnormal T cell 33 development, reduction of thymocyte numbers or altered T cells selection. Hence, ERK3 is 34 dispensable for pulmonary and T-cell functions. The perinatal lethality, lung and T-cell defects of 35 the previous ERK3 knockout mice are likely due to ERK3-unrelated effects of the inserted lacZ-36 neomycin-resistance-cassette. The knockout mouse of the closely related atypical MAPK 37 ERK4/MAPK4 is also normal suggesting redundant functions of both protein kinases. 38 Introduction (630 words) 39 40 Mitogen-activated protein kinase (MAPK) cascades are conserved eukaryote signaling modules 41 where the downstream effector kinases regulate cell proliferation, differentiation and cell death 42 by phosphorylation of protein substrates. MAPK are also regulators in many physiological 43 processes including development and immune response. Multiple MAPKs were described in 44 mammalian cells, which can be divided in five groups, the 'classical' mitogen-responsive MAPKs 45 (ERK1 and ERK2), the stress-activated JNKs (JNK1-3) and p38 MAPK (a,b,g,d), the big MAPK ERK5, 46 and the atypical MAPKs ERK3, ERK4 and ERK7 (1). MAPKs activity is classically regulated by dual 47 5) ERK3_1-target-seq.-FW: TGGACAGAGCACTGGAAG, 6) ERK3-loxP-RC: 131 CTTAAGACAGGAGTGTGGATC. 5+6 primer combination amplifies 943 bp of WT, 1055 bp of 132 exon3 floxed or 336 bp of exon3 deleted fragment. 133 PCR reactions were separated on 2% agarose gels and images acquired using INTAS Gel 134 documentation system. 135 136 137

ERK3 Is Involved in Regulating Cardiac Fibroblast Function

“…These mice were generated by inserting LacZ into exon 2. A more recent study using a conditional knockout where exon 3 of ERK3 was flanked by loxP observed that germline deletion of ERK3 resulted in ERK3-deficient mice that were viable (80).…”