Andreas Ehlich - Academia.edu (original) (raw)

Papers by Andreas Ehlich

Cell, Jan 1, 1993

The compartment of mouse B cell progenitors can be resolved into five developmentally related fra... more The compartment of mouse B cell progenitors can be resolved into five developmentally related fractions by multicolor flow cytometry. Using this system and employing mutant mice in which the membrane exon of the mu chain, the lambda 5 gene, or the JH locus was inactivated by gene targeting, we found that expression of the pre-B cell receptor complex is necessary for the transition from the large CD43+ to the small CD43- pre-B cell stage. We report the occurrence of immunoglobulin heavy and light chain gene rearrangement at the stage of large B cell precursors. We show that neither the pre-B cell receptor complex nor any gene rearrangement in the heavy chain locus is required for the induction of kappa light chain gene rearrangement in early B cell progenitors.

International Congress Series, 2003

Recent DNA typing methods (RFLP, STR, RAPD; e.g., [Naturwissenschaften 84 (1997)

Current Opinion in Immunology, 1995

The polymerase chain reaction allows the characterization of RNA and DNA sequences in samples as ... more The polymerase chain reaction allows the characterization of RNA and DNA sequences in samples as small as a single cell. The recent development of amplification systems designed to isolate rearranged immunoglobulin genes from single B lineage cells has provided a powerful tool to investigate various aspects of B-cell development.

Journal of Clinical Investigation, 2004

Parasympathetic slowing of the heart rate is predominantly mediated by acetylcholine-dependent ac... more Parasympathetic slowing of the heart rate is predominantly mediated by acetylcholine-dependent activation of the G protein-gated potassium (K + ) channel (I K,ACh ). This channel is composed of 2 inward-rectifier K + (Kir) channel subunits, Kir3.1 and Kir3.4, that display distinct functional properties. Here we show that subunit composition of I K,ACh changes during embryonic development. At early stages, I K,ACh is primarily formed by Kir3.1, while in late embryonic and adult cells, Kir3.4 is the predominant subunit. This change in subunit composition results in reduced rectification of I K,ACh , allowing for marked K + currents over the whole physiological voltage range. As a consequence, I K,ACh is able to generate the membrane hyperpolarization that underlies the strong negative chronotropy occurring in late-but not early-stage atrial cardiomyocytes upon application of muscarinic agonists. Both strong negative chronotropy and membrane hyperpolarization can be induced in early-stage cardiomyocytes by viral overexpression of the mildly rectifying Kir3.4 subunit. Thus, a switch in subunit composition is used to adopt I K,ACh to its functional role in adult cardiomyocytes.

Immunity, 1996

Allelic exclusion at the IgH locus was examined in B lineage cells of wild-type mice and mice una... more Allelic exclusion at the IgH locus was examined in B lineage cells of wild-type mice and mice unable to express the surrogate light chain molecule lambda 5 using a single-cell PCR approach. By analyzing B precursor cells containing two VHDHJH rearrangements, we found that in wild-type animals, cells are allelically excluded as soon as mu chains are expressed. Furthermore, we provide evidence that in cells expressing D mu proteins VH-->DHJH rearrangement is inhibited. In contrast, in the absence of lambda 5 protein, B precursor cells were allelically "included", indicating that allelic exclusion at the IgH locus requires expression of the pre-B cell receptor either containing a mu chain or a D mu chain. However, although mu chain double-producing B precursor cells are generated in lambda 5-deficient mice, such cells were not detected among surface immunoglobulin positive B cells.

The rearranged D23 V H D H J H gene segment was isolated by PCR amplification using a mixture of ... more The rearranged D23 V H D H J H gene segment was isolated by PCR amplification using a mixture of Taq and Pfu DNA polymerases and primer pair 23reX (5'-gga act cta gag aat ggc tgt ctt ggg gct gct c -3') -JH4A (Ehlich et al., 1994). To clone the promoters of the D23 heavy chain genes a "pan" -PCR genome walking strategy was employed (Siebert et al., 1995). Template library was prepared from genomic DNA isolated from the D23 hybridoma and digested with PvuII, EcoRV, SspI, DraI and ScaI restriction endonucleases prior to adapter ligation (Siebert et al., 1995). D23 IgH promoter was isolated with a gene specific primer PR1D23 (5'-gac aca gct tgg gaa tgt cac cag g-3') for the first round and a nested gene specific primer PR2D23 (5'-gga caa agc ttg gga atg tca cca ggc aga aga gc-3') for the second round. For further cloning, D23 VDJ and its promoter were amplified together from genomic DNA isolated from D23 hybridoma using primers specific for the upstream part of the cloned promoter Psp15 (5'-agt acc atg tcc cta agt ggc aag g-3') and JH4A (Ehlich et al., 1994). To introduce point mutations rendering D23 VDJ rearrangement nonproductive, we utilized PCR-mediated site directed mutagenesis using primers Mut1 (5' -caa gag cta agt ttc ttt aaa aat gaa c-3') and Mut2back (5' -gaa ttg tcc ttg ctg atg ctc agt ctg g-3') and the plasmid containing D23 VDJ and its promoter as a template. Resulting plasmids were sequenced.

Cell, 1993

Immunity, 2006

The immunoglobulin heavy chain repertoire is generated by somatic rearrangement of variable (V H ... more The immunoglobulin heavy chain repertoire is generated by somatic rearrangement of variable (V H ), diversity (D H ), and joining (J H ) elements. It can be further diversified by V H replacement, where nonrearranged V H genes invade preexisting V H D H J H joints. To study the impact and mechanism of V H replacement, we generated mice in which antibody production depends on the replacement of a nonproductive V H D H J H rearrangement inserted into its physiological position in the immunoglobulin heavy chain locus. In these mice a highly diverse heavy chain repertoire resulted from V H replacement and a second process of noncanonical V(D)J recombination, direct V H to J H joining. V H replacement rarely generated detectable sequence duplications but often proceeded through recombination between the conserved homologous sequences at the 3 0 end of V H . Thus, V H replacement is an efficient mechanism of antibody diversification, and its impact on the overall antibody repertoire could be greater than anticipated because it frequently leaves no molecular footprint.

Immunological Reviews, 1994

Whether a cell committed to the B-cell lineage finally succeeds in entering the pool of newly-gen... more Whether a cell committed to the B-cell lineage finally succeeds in entering the pool of newly-generated B cells depends on the Ig gene rearrangements it acquires during its differentiation. During the past decade, the immunoglobulin (Ig) gene rearrangement program has heen analyzed with the help of Abelson virus-transformed pre-B cell lines that spontaneously undergo rearrangement of Ig loci in vitro. Based on these studies, the following mode! of B-cell development was formulated (Alt et al. 1984, 1987): B-cell ...

Current Biology, 1994

Background: During B-cell development in the mouse, the V, DH and JH elements of the immunoglobul... more Background: During B-cell development in the mouse, the V, DH and JH elements of the immunoglobulin heavy chain (IgH) locus are rearranged, firstly by DH-JH joining, and then by VH-DHJH joining. In-frame ('productive') VHDHJH joints and DHJH joints in reading frame 2 (one of the three possible DH reading frames) allow the expression of F. and truncated Ip chains (DR proteins), respectively. The expression of such molecules from one of the two IgH loci of a cell is thought to interfere with VH-DJH recombination on the other IgH locus, and to guide the cells through further development.