Nucleocytoplasmic functions of the PDZ-LIM protein family: new insights into organ development

Abstract

Recent work on the PDZ-LIM protein family has revealed that it has important activities at the cellular level, mediating signals between the nucleus and the cytoskeleton, with significant impact on organ development. We review and integrate current knowledge about the PDZ-LIM protein family and propose a new functional role, sequestering nuclear factors in the cytoplasm. Characterized by their PDZ and LIM domains, the PDZ-LIM family is comprised of evolutionarily conserved proteins found throughout the animal kingdom, from worms to humans. Combining two functional domains in one protein, PDZ-LIM proteins have wide-ranging and multi-compartmental cell functions during development and homeostasis. In contrast, misregulation can lead to cancer formation and progression. New emerging roles include interactions with integrins, T-box transcription factors, and receptor tyrosine kinases. Facilitating the assembly of protein complexes, PDZ-LIM proteins can act as signal modulators, influence actin dynamics, regulate cell architecture, and control gene transcription.

Figure 1

The PDZ-LIM protein family. (A) General architecture and domain structure of the Eat-1/Tungus, Enigma and ALP subfamilies of PDZ-LIM proteins. The PDZ domain is indicated by the blue box and the LIM domain is depicted by the purple boxes. (B) Phylogenetic tree of members of the PDZ-LIM protein family. The single LIM domain (ALP subfamily) and three LIM domain (Enigma subfamily) gene structures evolved from a common ancestral gene encoding four LIM domains as found in the C. elegans Alp-1/Eat-1 and D. melanogaster Tungus proteins. While the ALP/Enigma subfamilies and LMO7 and LIM kinases appear to have a common ancestral origin (16,18), LMO7 and LIMK proteins reveal a complex evolution and are not included in the phylogenetic tree. Available sequences from C. elegans (Ce), zebrafish (Dr), chicken (Gg), mouse (Mm), and human (Hs) were compiled using MacVector 7.0 software (Oxford Molecular Company, Madison, WI). C. elegans Alp-1, the earliest known PDZ-LIM protein, was used as the out-group. The new nomenclature proposed by GenBank is displayed next to the phylogenetic tree with the previously published names on the brackets to the right. GenBank accession numbers for all genes discussed are as follows: C. elegans alp-1, CAE52903. D. melanogaster tungus, NM_137217; Ldb3: Homo sapiens, NM_007078; Mus Musculus, NM_011918; Gallus gallus, XM_421495; Danio rerio, DQ012157; D. melanogaster, NM_145757. Pdlim5: Homo sapiens, NM_006457; Mus Musculus, NM_019808; Gallus gallus, AJ851689; Danio rerio, BC045922. Pdlim7: Homo sapiens, NM_005451, Mus musculus, NM_026131: Gallus gallus, NM_001005345; Danio rerio, NM_200840. Pdlim1: Homo sapiens, NM_020992; Mus musculus, NM_016861; Gallus gallus, XM_426503; Danio rerio, BC092978. Pdlim3: Homo sapiens, NM_014476; Mus musculus, NM_016798; Gallus gallus, NM_001001764; Danio rerio, NM_001042718. Pdlim4: Homo sapiens, NM_003687; Mus musculus, NM_019417; Gallus gallus, NM_204839; Danio rerio, NM_001042696. Pdlim2: Homo sapiens, NM_176871; Mus musculus, NM_145978; Danio rerio, NM_001042766. Pdlim2 sequence could not be identified in the Gallus gallus genome.

Figure 2

PDZ-LIM proteins are required for muscle development and maintenance. High magnification example of zebrafish muscle phenotype after knock-down of pdlim7 with a splice-inhibiting morpholino (MO) antisense oligonucleotide (11). (A-B) Brightfield microscopy representing a lateral view of wild-type (A), and pdlim7 MO injected embryos (B) at 48 hpf. (C-D) Fluorescent microscopy showing a lateral view of wild-type (C), and pdlim7 MO injected embryos (D) at 48 hpf with F59 antibody (Developmental Studies Hybridoma Bank, University of Iowa) to detect slow muscle fibers. In comparison to the wild-type, the pdlim7 morphants exhibit elongated and severely disorganized slow muscle fibers. hpf=hours post-fertilization.

Figure 3

T-box transcription factors can localize outside the nucleus using a conserved nuclear export signal (NES). (A) Table listing reports of T-box proteins localized to the cytoplasm ranging from the mouse to the sponge. (B) Alignment of the NES from T-box proteins known to localize to the cytoplasm and their orthologs in the cephalochordate amphioxus and humans, demonstrating an evolutionarily conserved nuclear export mechanism. The NES consensus sequence was derived from proteins that utilize the CRM1 nuclear export pathway, where Φ represents hydrophobic residues (L, I, F, V, M, P) and X represents any amino acid. GenBank accession numbers for all sequences are as follows: Brachyury: Suberites domuncula AJ_544242; Branchiostoma floridae X_91903; Mus musculus NP_033335; Homo sapiens NM_003181. Tbx2: C. elegans Q_19691; Branchiostoma floridae (Tbx2/3) AF_262563; Homo sapiens NM_005994. Tbx5: Gallus gallus NM_204173; Branchiostoma floridae (Tbx4/5) AF_262564; Homo sapiens NM_000192. Tbr-1: Hemicentrotus pulcherrimus AB_048760; Branchiostoma floridae (Eomesdermin /Tbr1/Tbx21) AF_262568; Mus musculus NP_033348; Homo sapiens NM_006593. Eomesdermin: Danio rerio AF_329830; Homo sapiens NM_005442.

Figure 4

Model for the diverse and multi-compartmental functions of PDZ-LIM proteins. PDZ-LIM proteins are engaged in dynamic interactions with cellular proteins. For example, Pdlim7 facilitates shuttling of Tbx5 between nucleus and actin cytoskeleton. Export of Tbx5 from the nucleus is promoted by CRM1, and protein subcellular localization and function possibly regulated by reversible sumoylation. Besides tethering Tbx5 to the actin cytoskeleton, Pdlim7 can interact with other cellular proteins, including the activated InsR. Pdlim7 uses the same domain to bind either Tbx5 or InsR and this competitive interaction may provide a mechanism for Tbx5 release from filamentous actin and relocation to the nucleus. Transmembrane integrin receptors anchor the actin cytoskeleton to focal adhesions and transmit information from the extracellular environment. Several PDZ-LIM proteins have been localized to focal adhesions, providing the possibility that Pdlim7 could similarly engage with proteins clustered in focal adhesions including integrin receptors to sense extracellular stress.